Therapeutic compounds

ABSTRACT

The invention provides compounds of formula (I),  
                 
 
or a pharmaceutically acceptable salt of said compound, or a solvate of said compound or salt, wherein R 1 , R 2 , R 3 , HET, n, Q, X, Y, and Z are as described herein; compositions thereof; and uses thereof including treating Type 2 diabetes.

CROSS-REFERENCE TO RELATED APPLICATION

This patent application is a continuation of U.S. patent applicationSer. No. 11/129,277, filed May 12, 2005, which claims the benefit ofU.S. Provisional Application Ser. No. 60/664,305, filed on Mar. 21, 2005and U.S. Provisional Application Ser. No. 60/570,300, filed on May 12,2004, all of which are herein incorporated by reference in theirentirety.

FIELD OF THE INVENTION

The invention relates to selective inhibitors of the enzyme dipeptidylpeptidase-IV (DPP-IV), pharmaceutical compositions thereof, and usesthereof for treating diseases and conditions associated with proteinsthat are subject to processing by DPP-IV.

BACKGROUND OF THE INVENTION

DPP-IV (EC 3.4.14.5) is a serine protease that preferentially hydrolyzesan N-terminal dipeptide from proteins having proline or alanine in the2-position. DPP-IV is believed to be involved in diabetes, glucosetolerance, obesity, appetite regulation, lipidemia, osteoporosis,neuropeptide metabolism and T-cell activation, among others.Accordingly, administration of DPP-IV inhibitors in vivo preventsN-terminal degradation of substrate peptides, thereby resulting inhigher circulating concentrations of such peptides, and therapeuticbenefits associated with such elevated concentrations.

DPP-IV has been implicated in the control of glucose homeostasis becauseits substrates include the incretin peptides glucagon-like peptide 1(GLP-1) and gastric inhibitory polypeptide (GIP). Cleavage of theN-terminal amino acids from these peptides renders them functionallyinactive. GLP-1 has been shown to be an effective anti-diabetic therapyin Type 2 diabetic patients and to reduce the meal-related insulinrequirement in Type 1 diabetic patients. GLP-1 and/or GIP are believedto regulate satiety, lipidemia and osteogenesis. Exogenous GLP-1 hasbeen proposed as a treatment for patients suffering from acute coronarysyndrome, angina and ischemic heart disease.

Administration of DPP-IV inhibitors in vivo prevents N-terminaldegradation of GLP-1 and GIP, resulting in higher circulatingconcentrations of these peptides, increased insulin secretion andimproved glucose tolerance. On the basis of these observations, DPP-IVinhibitors are regarded as agents for the treatment of Type 2 diabetes,a disease in which glucose tolerance is impaired. In addition, treatmentwith DPP-IV inhibitors prevents degradation of Neuropeptide Y (NPY), apeptide associated with a variety of central nervous system disorders,and Peptide YY which has been linked to gastrointestinal conditions suchas ulcers, irritable bowel disease, and inflammatory bowel disease.

In spite of the early discovery of insulin and its subsequent widespreaduse in the treatment of diabetes, and the later discovery of and use ofsulfonylureas (e.g. chlorpropamide, tolbutamide, acetohexamide,biguanides (e.g., phenformin), metformin, thiazolidinediones (e.g.,rosiglitazone), and pioglitazone as oral hypoglycemic agents, thetreatment of diabetes remains less than satisfactory.

The use of insulin, necessary in Type 1 diabetic patients and about 10%of Type 2 diabetic patients in whom currently available oralhypoglycemic agents are ineffective, requires multiple daily doses,usually by self-injection. Determination of the appropriate dosage ofinsulin necessitates frequent estimations of the glucose concentrationin urine or blood. The administration of an excess dose of insulincauses hypoglycemia, with consequences ranging from mild abnormalitiesin blood glucose to coma, or even death.

Treatment of Type 2 diabetes usually comprises a combination of diet,exercise, oral agents, and in more severe cases, insulin. However, theclinically available hypoglycemics can have side effects that limittheir use. A continuing need for hypoglycemic agents, which may havefewer side effects or succeed where others fail, is clearly evident.

Poorly controlled hyperglycemia is a direct cause of the multiplicity ofcomplications (cataracts, neuropathy, nephropathy, retinopathy,cardiomyopathy) that characterize advanced Type 2 diabetes. In addition,Type 2 diabetes is a comorbid disease that frequently confoundshyperlipidemia, atherosclerosis and hypertension, adding significantlyto the overall morbidity and mortality attributable to those diseases.

Epidemiological evidence has firmly established hyperlipidemia as aprimary risk factor for cardiovascular disease (CVD) due toatherosclerosis. Atherosclerosis is recognized to be a leading cause ofdeath in the United States and Western Europe. CVD is especiallyprevalent among diabetic subjects, at least in part because of theexistence of multiple independent risk factors such as glucoseintolerance, left ventricular hypertrophy and hypertension in thispopulation. Successful treatment of hyperlipidemia in the generalpopulation, and in diabetic subjects in particular, is therefore ofexceptional medical importance.

Hypertension (high blood pressure) is a condition that can occur in manypatients in whom the causative agent or disorder is unknown. Such“essential” hypertension is often associated with disorders such asobesity, diabetes, and hypertriglyceridemia and it is known thathypertension is positively associated with heart failure, renal failure,and stroke. Hypertension can also contribute to the development ofatherosclerosis and coronary disease. Hypertension, together withinsulin resistance and hyperlipidemia, comprise the constellation ofsymptoms that characterize metabolic syndrome, also known as insulinresistance syndrome (IRS) and Syndrome X.

Obesity is a well-known and common risk factor for the development ofatherosclerosis, hypertension, and diabetes. The incidence of obesityand its related sequelae is increasing worldwide. Currently, fewpharmacological agents are available that reduce adiposity effectivelyand acceptably.

Osteoporosis is a progressive systemic disease characterized by low bonedensity and microarchitectural deterioration of bone tissue, with aconsequent increase in bone fragility and susceptibility to fracture.Osteoporosis and the consequences of compromised bone strength are asignificant cause of frailty, and of increased morbidity and mortality.

Heart disease is a major health problem throughout the world. Myocardialinfarctions are a significant source of mortality among thoseindividuals with heart disease. Acute coronary syndrome denotes patientswho have or are at high risk of developing an acute myocardialinfarction (MI).

Though there are therapies available for the treatment of diabetes,hyperglycemia, hyperlipidemia, hypertension, obesity, and osteoporosisthere is a continuing need for alternative and improved therapies.

Various indications for DPP-IV inhibitors are discussed in Augustyns, etal., Curr. Medicinal Chem., 6, 311 (1999); Ohnuki, et al., Drugs of theFuture, 1999, 24, 665-670 (1999); Villhauer, et al., Annual Reports inMedicinal Chemistry, 36, 191-200 (2001); Drucker, Expert Opin. Invest.Drugs, 12, 87-100 (2003); and Weideman, et al., Curr. Opin. Invest.Drugs, 4, 412-420 (2003).

Orally administered compounds that inhibit DPP-IV have recently beenprepared, such as those disclosed in International Application WO02/14271.

DPP-IV inhibitors, such as those disclosed in WO 02/14271, are believedto act by inhibiting the degradation of the natural hormones, GLP-1 andGIP. Therefore, it is important that a suitable concentration of theDPP-IV inhibitor be available in plasma to inhibit DPP-IV coincidentlywith the secretion of these GLP-1 and GIP hormones. To achieve suchplasma concentrations, it is preferred that the DPP-IV inhibitorcompounds maintain a higher plasma concentration over time than thatwhich would be expected for other DPP-IV inhibitor compounds, such asthose disclosed in WO 02/14271.

Therefore, what is needed is an orally administered DPP-IV inhibitorcompound that has equivalent or better DPP-IV inhibitory activity andthat maintains a higher plasma concentration over time.

SUMMARY OF THE INVENTION

The present invention relates to compounds having the structure ofFormula (I)

or a pharmaceutically acceptable salt of said compound, or a solvate ofsaid compound or salt, wherein:

R¹ is —(C₁-C₆)alkyl, —(C₁-C₆)alkoxy, —(C₁-C₆)arylalkyl, —NR^(a)R^(b),hydroxy, cyano, aryl, or heteroaryl, wherein said —(C₁-C₆)alkyl, saidaryl, or said heteroaryl is optionally substituted independently withone to three —COOH, —C(O)(C₁-C₆)alkoxy, —C(O)(C₁-C₆)alkyl,—C(O)NR^(a)R^(b), cyano, halogen, nitro, trifluoromethyl, —(C₁-C₆)alkyl,—(C₁-C₆)alkoxy, —(C₃-C₆)cycloalkyl, or phenyl, and wherein R^(a) andR^(b) are, independently, hydrogen, —(C₁-C₆)alkyl, aryl, or heteroaryl,or R^(a) and R^(b), taken together with the nitrogen atom to which theyare attached, form a four- to six-membered heterocyclic ring, whereinsaid ring optionally incorporates an additional one or two nitrogen,oxygen, or sulfur ring heteroatoms;

R² and R³ are, independently, hydrogen, halogen, —(C₁-C₆)alkyl, or—(C₃-C₈)cycloalkyl;

Q is a covalent bond, —C(O)—, or —SO₂—;

HET is a heterocycloalkyl ring moiety, optionally substituted with: (A)one to four —(C₁-C₆)alkyl, optionally substituted with one to sixhalogen atoms, —(C₁-C₆)alkoxy, cyano, halogen, hydroxy, or —NR^(a)R^(b),or (B) —(C₁-C₆)arylalkyl, optionally substituted with one to six halogenatoms, —(C₁-C₆)alkoxy, cyano, halogen, hydroxy, or —NR^(a)R^(b);

n is zero or one;

X is —CH₂—, —CHF—, or —CF₂— and Y is —CH₂—, —CHF—, or —CF₂—, providedthat when n is one X and Y are not both CH₂ and when n is zero X is—CH₂—; and

Z is hydrogen or cyano.

The present invention also relates to a pharmaceutical compositioncomprising a therapeutically effective amount of a compound of thepresent invention, or a pharmaceutically acceptable salt of thecompound, or a solvate of the compound or salt, and a pharmaceuticallyacceptable carrier, vehicle, diluent or excipient.

The present invention further relates to a method of treating diabetescomprising administering to a mammal in need of such treatment atherapeutically effective amount of a compound of the present invention,or a pharmaceutically acceptable salt of the compound, or a solvate ofthe compound or salt. Preferably, the type of diabetes treated is Type 2diabetes.

The present invention additionally relates to a method of treating acondition mediated by dipeptidyl peptidase-IV in a mammal comprisingadministering to said mammal in need of such treatment a therapeuticallyeffective amount of a compound of the present invention, or apharmaceutically acceptable salt of said compound, or a solvate of saidcompound or salt.

The compounds, salts, solvates and pharmaceutical compositions of thepresent invention are useful for the treatment of Type 2 diabetes, Type1 diabetes, impaired glucose tolerance, hyperglycemia, metabolicsyndrome (also known as Syndrome X or insulin resistance syndrome), anddiabetic complications such as sugar cataracts, diabetic neuropathy,diabetic nephropathy, diabetic retinopathy and diabetic cardiomyopathy;and also the prevention or mitigation of disease progression in Type 1and Type 2 diabetes.

The compounds, salts, solvates and pharmaceutical compositions of thepresent invention are also useful for the treatment of diabetes-relatedatherosclerosis; obesity, conditions exacerbated by obesity,hypertension, hyperlipidemia, osteoporosis, osteopenia, frailty, boneloss, bone fracture, acute coronary syndrome, infertility due topolycystic ovary syndrome, short stature due to growth hormonedeficiency, anxiety, depression, insomnia, chronic fatigue, epilepsy,eating disorders, chronic pain, alcohol addiction, diseases associatedwith intestinal motility, ulcers, irritable bowel syndrome, inflammatorybowel syndrome, short bowel syndrome, and cancer.

DETAILED DESCRIPTION

The terms used to describe the present invention have the followingmeanings herein.

The phrase “pharmaceutically acceptable” indicates that the designatedcarrier, vehicle, diluent, excipient(s), and/or salt is generallychemically and/or physically compatible with the other ingredientscomprising the formulation, and physiologically compatible with therecipient thereof.

The carbon atom content of the various hydrocarbon-containing moietiesherein may be indicated by a prefix designating the minimum and maximumnumber of carbon atoms in the moiety, for example, the prefixes(C_(a)-C_(b))alkyl, and C_(a-b)alkyl, indicate an alkyl moiety of theinteger “a” to “b” carbon atoms, inclusive. Thus, for example,(C₁-C₆)alkyl and C₁₋₆-alkyl refer to an alkyl group of one to six carbonatoms inclusive.

The term “alkyl” denotes a straight or branched chain of carbon atoms,wherein the alkyl group optionally incorporates one or more double ortriple bonds, or a combination of double bonds and triple bonds.Examples of alkyl groups include methyl, ethyl, propyl, isopropyl,butyl, isobutyl, vinyl, allyl, 2-methylpropenyl, 2-butenyl,1,3-butadienyl, ethynyl, propargyl, and the like.

The term “alkoxy” refers to straight or branched, monovalent, saturatedaliphatic chains of carbon atoms bonded to an oxygen atom that isattached to a core structure. Examples of alkoxy groups include methoxy,ethoxy, propoxy, butoxy, iso-butoxy, tert-butoxy, and the like.

The term “cycloalkyl” denotes a saturated monocyclic or bicycliccycloalkyl group. Cycloalkyl groups may be optionally fused to aromatichydrocarbons such as benzene to form fused cycloalkyl groups, such asindanyl and the like. Examples of cycloalkyl groups include cyclopropyl,cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and the like.

The term “halogen” or “halo” represents chloro, bromo, fluoro, and iodoatoms and substituents.

The term “aryl” denotes a monocyclic or polycyclic aromatic hydrocarbongroup, for example, anthracenyl, fluorenyl, naphthyl, phenanthrenyl,phenyl, and the like.

The term “arylalkyl” means an alkyl group, as defined hereinabove,wherein at least one of the hydrogen atoms thereof has been substitutedwith an aryl group, also as defined hereinabove. Examples of arylalkylgroups include, inter alia, benzyl groups.

The term “heterocycloalkyl”, as employed with reference to HEThereinabove, refers to a saturated four- to eight-membered heterocyclicring system, optionally fused to a five- or six-membered aromatic orheteroaromatic ring system. Examples of heterocycloalkyl groups comprisehomopiperazinyl, piperazinyl, piperidyl, pyrrolidinyl, azetidinyl,2-aza-bicyclo[2.2.1]heptanyl, 3-aza-bicyclo[3.1.0]hexanyl,2,5-diaza-bicyclo[2.2.1]heptanyl,5,6,7,8-tetrahydro-2H-imidazo[1,2-a]pyrazinyl,5,6,7,8-tetrahydro[1,2,4]triazolo[4,3-a]pyrazinyl,4,5,6,7-tetrahydropyrazolo[1,5-a]pyrazinyl,5,6,7,8-tetrahydropyrido[3,4-d]pyrimidyl,5,6,7,8-tetrahydropyrido[4,3-d]pyrimidyl,octahydropyrrolo[3,4-b]pyrrolyl, octahydropyrrolo[3,4-c]pyrrolyl,6-azabicyclo[3.2.1]octanyl, 3,8-diazabicyclo[3.2.1]octanyl,2,3-dihydrospiro[indene-1,4′-piperidyl]spiro[indene-1,4′-piperidyl],1-oxa-8-azaspiro[4.5]decanyl, 8-azabicyclo[3.2.1]octanyl,2,3,4,5-tetrahydrobenzo[f][1,4]oxazepinyl,hexahydro-2H-pyrrolo[3,4-d]isothiazolyl-1,1-dioxide,2,7-diazaspiro[4.4]nonanyl,6,7,8,9-tetrahydro-5H-[1,2,4]triazolo[4,3-g][1,4]diazepinyl,5,6-dihydro-8H-imidazo[1,2-a]pyrazinyl,5,6-dihydro-8H-[1,2,4]triazolo[4,3-a]pyrazinyl,7,8-dihydro-5H-pyrido[4,3-a]pyrimidyl,7,8-dihydro-5H-pyrido[4,3-d]pyrimidyl, pyrazolo[1,5-a]pyrimidyl, and thelike.

The term “heteroaryl” denotes a monocyclic or polycyclic aromaticheterocyclic ring system. Examples of heteroaryl groups comprisebenzoisothiazolyl, benzisoxazolyl, benzooxazolyl, benzothiazolyl,benzofuranyl, benzothienyl, benzimidazolyl, cinnolinyl, furanyl,furopyridyl, imidazolopyrimidyl, imidazolyl, indazolyl, indolyl,isoquinolyl, isothiazolyl, isoxadiazolyl, isoxazolyl, oxazolopyridyl,oxadiazolyl, oxazolyl, phthalazinyl, pteridinyl, pyrazinyl, pyridazinyl,pyrrolopyrimidyl, pyrrolopyridyl, pyrazolopyrimidyl, pyrazolyl, pyridyl,pyrimidyl, pyrrolyl, quinazolyl, quinolyl, quinoxalinyl, tetrazolyl,thiazolyl, thiadiazolyl, thiazolopyridyl, thienopyridyl, thienyl,triazinyl, triazolyl, 1,1-dioxo-1H-1,2-benzoisothiazolyl,oxazolopyridyl, and the like.

The term “oxo”, means a carbonyl group formed by the combination of acarbon atom and an oxygen atom.

The term “substituted” means that a hydrogen atom on a molecule has beenreplaced with a different atom or molecule. The atom or moleculereplacing the hydrogen atom is denoted as a “substituent.”

The symbol “-” represents a covalent bond.

The phrase “inert solvent” refers to a solvent, or mixture of solvents,that does not interact with starting materials, reagents, intermediates,or products in a manner that adversely affects their desired properties.

The terms “treating”, “treated”, or “treatment” as employed hereinincludes preventative (e.g., prophylactic), palliative, and curativeuses or results.

The phrase “therapeutically effective amount” means an amount of acompound of the present invention that (i) treats or prevents theparticular disease, condition, or disorder, (ii) attenuates,ameliorates, or eliminates one or more symptoms of the particulardisease, condition, or disorder, or (iii) prevents or delays the onsetof one or more symptoms of the particular disease, condition, ordisorder described herein.

The term “mammal” is an individual animal that is a member of thetaxonomic class Mammalia. The class Mammalia includes, for example,humans, monkeys, chimpanzees, gorillas, cattle, swine, horses, sheep,dogs, cats, mice and rats. In the present invention, the preferredmammal is a human.

Preferably, the compounds of the present invention have the structure ofFormula (I) wherein:

R¹ is aryl or heteroaryl, optionally substituted independently with oneto three cyano, halogen, nitro, trifluoromethyl, —(C₁-C₆)alkyl,—(C₁-C₆)alkoxy, —(C₃-C₆)cycloalkyl, or phenyl;

R² is —H or —(C₁-C₆)alkyl;

R³ is —H—(C₁-C₆)alkyl; and

HET is azetidinyl, piperazinyl, piperidinyl, pyrrolidinyl,5,6-dihydro-8H-imidazo[1,2-a]pyrazin-7-yl,5,6-dihydro-8H-[1,2,4]triazolo[4,3-a]pyrazin-7-yl, or7,8-dihydro-5H-pyrido[4,3-a]pyrimidin-6-yl.

More preferably, the compounds of the present invention have thestructure of Formula (IA)

wherein R¹ is benzoisothiazolyl, benzisoxazolyl, isothiazolyl,isoxazolyl, oxazolopyridyl, pyrazinyl, pyridinyl, pyrimidinyl,quinolinyl, quinoxalinyl, thiadiazolyl, triazinyl, or1,1-dioxo-1H-1,2-benzoisothiazolyl.

In the present invention, it is preferred, for the compounds of Formula(IA), that R¹ is pyridinyl or pyrimidinyl and more preferred that R¹ ispyridinyl or pyrimidinyl, n is 1, X is —CF₂— and Y is —CH₂—.

In the present invention, the compound(3,3-difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(pyrimidin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)methanone,or a pharmaceutically acceptable salt of said compound is mostpreferred.

In an alternate embodiment, a compound selected from the groupconsisting of:

((2S,4S)-4-(3-(trifluoromethyl)-5,6-dihydro-[1,2,4]triazolo[4,3-a]pyrazin-7(8H)-yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone,

(3,3-difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(oxazolo[5,4-b]pyridin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone,

(3,3-difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(4-methylpyrimidin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone,

((2S,4S)-4-(2-(trifluoromethyl)-7,8-dihydropyrido[4,3-d]pyrimidin-6(5H)-yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone,

((S)-3-fluoro-pyrrolidin-1-yl)-{(2S,4S)-4-[4-(3-trifluoromethyl-pyridin-2-yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone,

((S)-3-fluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-trifluoromethyl-7,8-dihydro-5H-pyrido[4,3d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone,

(3,3-difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[4,5-c]pyridin-2-yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone,

[(2S,4S)-4-(2-cyclopropyl-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-(3-fluoro-azetidin-1-yl)-methanone,

(3,3-difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-ethoxy-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone,

2-{4-[(3S,5S)-5-(3-fluoro-azetidine-1-carbonyl)-pyrrolidin-3-yl]-piperazin-1-yl}-nicotinonitrile,

((S)-3-fluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-b]pyridin-2-yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone,

(3-fluoro-azetidin-1-yl)-[(2S,4S)-4-(2-trifluoromethyl-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone,

2-{4-[(3S,5S)-5-((S)-3-fluoro-pyrrolidine-1-carbonyl)-pyrrolidin-3-yl]-piperazin-1-yl}-nicotinonitrile,

(3-fluoro-azetidin-1-yl)-{(2S,4S)-4-[4-(2-trifluoromethyl-quinolin-4-yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone,

((3R*,4S*)-3,4-difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-trifluoromethyl-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone,and

((3R*,4S*)-3,4-difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-b]pyridin-2-yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone,or a pharmaceutically acceptable salt of said compound, is preferred.

The compounds of the present invention contain all contain at least twostereogenic centers, specifically the (2S,4S)pyrrolidin-2-yl,stereogenic centers shown below in Formula (I).

The compounds of the present invention may be resolved into the pureenantiomers by methods known to those skilled in the art, for example byformation of diastereoisomeric salts which may be separated, forexample, by crystallization; formation of diastereoisomeric derivativesor complexes which may be separated, for example, by crystallization,gas-liquid or liquid chromatography; selective reaction of oneenantiomer with an enantiomer-specific reagent, for example enzymaticesterification; or gas-liquid or liquid chromatography in a chiralenvironment, for example on a chiral support for example silica with abound chiral ligand or in the presence of a chiral solvent. It will beappreciated that where the desired stereoisomer is converted intoanother chemical entity by one of the separation procedures describedabove, a further step is required to liberate the desired enantiomericform. Alternatively, the specific stereoisomers may be synthesized byusing an optically active starting material, by asymmetric synthesisusing optically active reagents, substrates, catalysts or solvents, orby converting one stereoisomer into the other by asymmetrictransformation.

Wherein said compounds contain one or more additional stereogeniccenters, those skilled in the art will appreciate that alldiastereoisomers and diastereoisomeric mixtures of the compoundsillustrated and discussed herein are within the scope of the presentinvention. These diastereoisomers may be isolated by methods known tothose skilled in the art, for example, by crystallization, gas-liquid orliquid chromatography. Altenatively, intermediates in the course of thesynthesis may exist as racemic mixtures and be subjected to resolutionby methods known to those skilled in the art, for example by formationof diastereoisomeric salts which may be separated, for example, bycrystallization; formation of diastereoisomeric derivatives or complexeswhich may be separated, for example, by crystallization, gas-liquid orliquid chromatography; selective reaction of one enantiomer with anenantiomer-specific reagent, for example enzymatic esterification; orgas-liquid or liquid chromatography in a chiral environment, for exampleon a chiral support for example silica with a bound chiral ligand or inthe presence of a chiral solvent. It will be appreciated that where thedesired stereoisomer is converted into another chemical entity by one ofthe separation procedures described above, a further step is required toliberate the desired enantiomeric form. Alternatively, specificstereoisomers may be synthesized by asymmetric synthesis using opticallyactive reagents, substrates, catalysts or solvents, or by converting onestereoisomer into the other by asymmetric transformation.

Certain compounds of Formula (I) may exist in different stableconformational forms which may be separable. Torsional asymmetry due torestricted rotation about an asymmetric single bond, for example becauseof steric hindrance or ring strain, may permit separation of differentconformers. The present invention includes each conformational isomer ofcompounds of Formula (I) and mixtures thereof. Practitioners willappreciate that certain compounds of Formula (I) may exist in tautomericform, i.e., that an equilibrium exists between two isomers which are inrapid equilibrium with each other. A common example of tautomerism isketo-enol tautomerism, i.e.,

Examples of such compounds of the present invention include, inter alia,hydroxypyridines (pyridones) and hydroxypyrmidines (pyridones). Inparticular, a person skilled in the art will recognize that ahydroxypyridine of the instant invention can exist as two separatetautomers, e.g.,

The degree to which one tautomer is present over the other depends uponvarious factors, including substitution pattern and solvent type. Otherexamples in accordance with the present invention will be recognized bythose skilled in the art. All tautomeric forms of Formula (I) areincluded within the scope of the claimed invention.

The compounds of the present invention may exist in unsolvated as wellas solvated forms with pharmaceutically acceptable solvents such aswater, ethanol, and the like, and it is intended that the inventionembrace unsolvated forms, solvated forms and mixtures of solvated forms.

Certain compounds of Formula (I) and their salts and solvates may existin more than one crystal form. Polymorphs of compounds represented byFormula (I) form part of this invention and may be prepared bycrystallization of a compound of Formula (I) under different conditions.For example, using different solvents or different solvent mixtures forrecrystallization; crystallization at different temperatures; variousmodes of cooling, ranging from very fast to very slow cooling duringcrystallization. Polymorphs may also be obtained by heating or melting acompound of Formula (I) followed by gradual or fast cooling. Thepresence of polymorphs may be determined by solid probe nmrspectroscopy, ir spectroscopy, differential scanning calorimetry, powderX-ray diffraction or such other techniques.

This invention also includes isotopically-labeled compounds, which areidentical to those described by Formula (I), but for the fact that oneor more atoms are replaced by an atom having an atomic mass or massnumber different from the atomic mass or mass number usually found innature. Examples of isotopes that can be incorporated into compounds ofthe invention include isotopes of hydrogen, carbon, nitrogen, oxygen,sulfur and fluorine, such as ²H, ³H, ¹³C, ¹⁴C, ¹⁵N, ¹⁸O, ¹⁷O, ³⁵S, ³⁶Cl,¹²⁵I, ¹²⁹I and ¹⁸F respectively. Compounds of the present invention,prodrugs thereof, and pharmaceutically acceptable salts of the compoundsor of the prodrugs which contain the aforementioned isotopes and/orother isotopes of other atoms are within the scope of this invention.Certain isotopically-labeled compounds of the present invention, forexample those into which radioactive isotopes such as ³H and ¹⁴C areincorporated, are useful in drug and/or substrate tissue distributionassays. Tritiated (i.e., ³H), and carbon-14 (i.e., ¹⁴C), isotopes areparticularly preferred for their ease of preparation and detectability.Further, substitution with heavier isotopes such as deuterium (i.e.,²H), can afford certain therapeutic advantages resulting from greatermetabolic stability, for example increased in vivo half-life or reduceddosage requirements and, hence, may be preferred in some circumstances.Isotopically labeled compounds of Formula (I) of this invention andprodrugs thereof can generally be prepared by carrying out theprocedures disclosed in the schemes and/or in the Examples below, bysubstituting a readily available isotopically labeled reagent for anon-isotopically labeled reagent.

Pharmaceutically acceptable salts, as used herein in relation tocompounds of the present invention, include pharmaceutically acceptableinorganic and organic salts of said compound. These salts can beprepared in situ during the final isolation and purification of acompound, or by separately reacting the compound or prodrug with asuitable organic or inorganic acid and isolating the salt thus formed.Representative salts include, but are not limited to, the hydrobromide,hydrochloride, hydroiodide, sulfate, bisulfate, nitrate, acetate,trifluoroacetate, oxalate, besylate, palmitate, pamoate, malonate,stearate, laurate, malate, borate, benzoate, lactate, phosphate,hexafluorophosphate, benzene sulfonate, tosylate, formate, citrate,maleate, fumarate, succinate, tartrate, naphthylate, mesylate,glucoheptonate, lactobionate and laurylsulphonate salts, and the like.These may also include cations based on the alkali and alkaline earthmetals, such as sodium, lithium, potassium, calcium, magnesium, and thelike, as well as non-toxic ammonium, quaternary ammonium, and aminecations including, but not limited to, ammonium, tetramethylammonium,tetraethylammonium, methylamine, dimethylamine, trimethylamine,triethylamine, ethylamine, and the like. For additional examples see,for example, Berge, et al., J. Pharm. Sci., 66, 1-19 (1977).

The compounds of the present invention may be isolated and used per seor in the form of their pharmaceutically acceptable salts or solvates.In accordance with the present invention, compounds with multiple basicnitrogen atoms can form salts with varying number of equivalents ofacid. It will be understood by practitioners that all such salts arewithin the scope of the present invention.

A prodrug of a compound of Formula (I) may be one formed in aconventional manner with a functional group of the compound, such aswith an amino, hydroxy or carboxy group. The term “prodrug” means acompound that is transformed in vivo to yield a compound of Formula (I)or a pharmaceutically acceptable salt or solvate of the compound. Thetransformation may occur by various mechanisms, such as throughhydrolysis in blood. A discussion of the use of prodrugs is provided byT. Higuchi and W. Stella, “Pro-drugs as Novel Delivery Systems,” Vol. 14of the A.C.S. Symposium Series, and in Bioreversible Carriers in DrugDesign, ed. Edward B. Roche, American Pharmaceutical Association andPergamon Press, 1987.

For example, if a compound of the present invention incorporates anamine functional group, a prodrug can be formed by the replacement of ahydrogen atom in the amine group with a group such as R-carbonyl,RO-carbonyl, NRR′-carbonyl where R and R′ are each independently(C₁-C₁₀)alkyl, (C₃-C₇)cycloalkyl, benzyl, or R-carbonyl is a naturalα-aminoacyl or natural α-aminoacyl-natural α-aminoacyl, —C(OH)C(O)OY′wherein Y′ is H, (C₁-C₆)alkyl or benzyl, —C(OY₀)Y₁ wherein Y₀ is(C₁-C₄)alkyl and Y₁ is (C₁-C₆)alkyl, carboxy(C₁-C₆)alkyl,amino(C₁-C₄)alkyl or mono-N— or di-N,N—(C₁-C₆)alkylaminoalkyl, —C(Y₂)Y₃wherein Y₂ is H or methyl and Y₃ is mono-N— or di-N,N—(C₁-C₆)alkylamino,morpholino, piperidin-1-yl or pyrrolidin-1-yl.

Similarly, if a compound of the present invention contains an alcoholfunctional group, a prodrug can be formed by the replacement of thehydrogen atom of the alcohol group with a group such as(C₁-C₆)alkanoyloxymethyl, 1-((C₁-C₆)alkanoyloxy)ethyl,1-methyl-1-((C₁-C₆)alkanoyloxy)ethyl, (C₁-C₆)alkoxycarbonyloxymethyl,N—(C₁-C₆)alkoxycarbonylaminomethyl, succinoyl, (C₁-C₆)alkanoyl,α-amino(C₁-C₄)alkanoyl, arylacyl and α-aminoacyl, orα-aminoacyl-α-aminoacyl, where each α-aminoacyl group is independentlyselected from the naturally occurring L-amino acids, P(O)(OH)₂,—P(O)(O(C₁-C₆)alkyl)₂ or glycosyl (the radical resulting from theremoval of a hydroxyl group of the hemiacetal form of a carbohydrate).

If a compound of the present invention contains a carboxylic acidfunctional group, a prodrug can comprise an ester formed by thereplacement of the hydrogen atom of the acid group with a group such as(C₁-C₈)alkyl, (C₂-C₁₂)alkanoyloxymethyl, 1-(alkanoyloxy)ethyl havingfrom 4 to 9 carbon atoms, 1-methyl-1-(alkanoyloxy)-ethyl having from 5to 10 carbon atoms, alkoxycarbonyloxymethyl having from 3 to 6 carbonatoms, 1-(alkoxycarbonyloxy)ethyl having from 4 to 7 carbon atoms,1-methyl-1-(alkoxycarbonyloxy)ethyl having from 5 to 8 carbon atoms,N-(alkoxycarbonyl)aminomethyl having from 3 to 9 carbon atoms,1-(N-(alkoxycarbonyl)amino)ethyl having from 4 to 10 carbon atoms,3-phthalidyl, 4-crotonolactonyl, gamma-butyrolacton-4-yl,di-N,N—(C₁-C₂)alkylamino(C₂-C₃)alkyl (such as β-dimethylaminoethyl),carbamoyl-(C₁-C₂)alkyl, N,N-di(C₁-C₂)alkylcarbamoyl-(C₁-C₂)alkyl andpiperidino-, pyrrolidino- or morpholino(C₂-C₃)alkyl.

In general, the compounds of Formula (I) of this invention may beprepared by methods that include processes known in the chemical arts,particularly in light of the description contained herein. Certainprocesses for the manufacture of the compounds of Formula (I) of thisinvention are illustrated by the following reaction schemes. Otherprocesses are described in the experimental section. The methodsdisclosed in the instant Schemes and Examples are are intended forpurposes of exemplifying the instant invention, and are not to beconstrued in any manner as limitations thereon.

Some of the starting compounds for the reactions described in theschemes and Examples are prepared as illustrated herein. All otherstarting compounds may be obtained from general commercial sources, suchas Sigma-Aldrich Corporation, St. Louis, Mo.

In the discussions below, the following abbreviations are used: BOC(tert-butoxycarbonyl), Cbz (benzyloxycarbonyl), DMF(N,N-dimethylformamide), NMP (N-methyl-2-pyrrolidinone), DMAC(N,N-dimethylacetamide), DME (dimethoxyethane), DMSO(dimethylsulfoxide), EtOAC (ethyl acetate), EtOH (ethanol), MeOH(methanol), TFA (trifluoroacetic acid), TFAA (trifluoroaceticanhydride), TEA (triethylamine), THF (tetrahydrofuran), DIPEA(diisopropylethylamine), EDC(1-(3-dimethylaminopropyl)-3-carbodiimide)), DCC(dicyclohexylcarbodiimide), CDI (1,1′-carbonyldiimidazole), HATU(O-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate), HOAT (1-hydroxy-7-azabenzotriazole), HOBT(N-hydroxybenzotriazole), and EEDQ(2-ethoxy-1-ethoxycarbonyl-1,2-dihydroquinoline).

A generalized method for preparing the compounds of formula (I) isdepicted in Scheme 1 hereinbelow.

In Scheme 1, a compound of formula (II), prepared as described in Scheme2, wherein P represents a nitrogen-protecting group, is deprotectedaccording to known methods. If P represents BOC, deprotection istypically effected by first treating (II), dissolved in a solvent suchas EtOAc, ether dioxane or water, with optional cooling at a suitabletemperature, such as about 0° C., with acid (e.g., hydrogen chloride)for a suitable time, such as about 5 minutes to about an hour. Thesolution is allowed to warm to room temperature (RT), followed bystirring for an additional amount of time, typically an additional 30minutes to about 16 hours. Preferably, the reaction mixture is stirredabout 15 minutes, allowed to reach room temperature, then stirred anadditional 30 minutes. Alternatively, (II) is dissolved in TFA and,after a suitable time (e.g., about 30 min to about 24 hours), excess TFAis removed in vacuo, and the residual product is triturated with asolvent such as ether. If P represents Cbz, deprotection may beperformed by hydrogenolysis in the presence of catalyst, such as 10%palladium or palladium hydroxide, in a suitable solvent such as EtOH orEtOAC at a pressure of about 30 psi to about 60 psi, for a sufficientperiod of time, usually overnight, at a temperature of between about 20°C. and about 80° C. Preferably, hydrogenolysis is effected at a pressureof about 45 psi at room temperature.

The compounds of formula (II) may be prepared by coupling anappropriately-substituted carboxylic acid derivative (III) with anappropriately-substituted amine derivative (IV) as depicted hereinbelowin Scheme 2.

The coupling is typically accomplished by combining (Ill) and (IV) in areaction-inert solvent, preferably an aprotic solvent such asacetonitrile, dichloromethane, DMF, THF, or chloroform. A couplingagent, such as EDC, HATU, DCC, EEDQ, CDI, pivaloyl chloride ordiethylphosphorylcyanide is then added, optionally in the presence of abase, such as TEA or pyridine, and an optional adjuvant, such as HOBT orHOAT. The coupling is typically effected at a temperature of betweenabout 0° C. and about 50° C., for a suitable time, such as from aboutone hour and about 24 hours, for example about 16 hours. For adiscussion of other conditions useful for coupling carboxylic acids seeHouben-Weyl, Vol. XV, Part II, E. Wunsch, Ed., G. Theime Verlag, (1974),Stuttgart; M. Bodansky, “Principles of Peptide Synthesis”,Springer-Verlag Berlin (1984); and “The Peptides: Analysis, Synthesisand Biology” (ed. E. Gross and J. Meienhofer), Vols. 1-5 (Academic PressNY 1979-1983). The compounds of formulae (Ill) and (IV) may be preparedby known methods or, alternatively, according to the exemplarypreparative procedures described hereinbelow. For exemplary preparationsof formula (IV) amines, see PCT International Application PublicationNo. WO 2003/101958 and U.S. Pat. No. 6,710,040, the disclosure of whichis incorporated herein by reference.

Alternatively, the compounds of formula (II) may be prepared asdescribed below in Scheme 3.

In Scheme 3, the compounds of formula (II) are prepared by reductiveamination of a protected ketone (V), prepared as described hereinbelowin Scheme 4, with an appropriately-substituted heterocycloalkylamine(VI). Such amination reactions are well-known to one skilled in the art.See, for example, A. F. Abdel-Magid, et al., J. Org. Chem., 61, 3849(1996); R. F. Borch, et al., J. Am. Chem. Soc., 93, 2897 (1971); and S.Bhattacharyya, et al., Synlett, 1079 (1995). The formula (VI) amines arewell-known in the relevant art and may be obtained commercially orprepared by known methods. See, for example, D. A. Horton, et al., Chem.Rev., 103, 893-930 (2003), H. Fukui, et al, Heterocycles, 56, 257-264(2002), M. Y. Chu-Moyer, et al., J. Org. Chem., 60, 5721-5725 (1995),and J. P. Yevich, et al., J. Med. Chem., 29, 359-369 (1986).

Typically, (V) and (VI) are condensed in the presence of a reducingagent such as sodium borohydride, sodium cyanoborohydride, sodiumtriacetoxyborohydride, tetramethylammonium triacetoxyborohydride, orhydrogen in the presence of a catalyst (10% Pd/C, platinum oxide, etc.),optionally in the presence of an acid (e.g. acetic acid (AcOH),hydrochloric acid, etc.). The coupling is normally effected in areaction-inert solvent, such as 1,2-dichloroethane, THF, DMF, EtOH, orMeOH. The reaction is performed at a suitable temperature, such as 0 to50° C., for a suitable period of time, such as between about one toabout 24 hours, for example, about 16 hours.

The compounds of formula (V) may be prepared as described hereinbelow inScheme 4, beginning with, as appropriate, commercially availablecarboxylic acid (VII), ketocarboxylic acid (IX), or ketoester (X).

In Scheme 4, Step 1, protected acid (VII) is coupled with amine (IV) asdescribed hereinabove in Scheme 2 to afford alcohol (VIII).

In Scheme 4, Step 2, alcohol (VIII) is oxidized to ketone (Va) bytreating (VIII) with an oxidizing agent in a reaction-inert solvent.Examples of appropriate oxidizing agents comprise pyridine/sulfurtrioxide in DMSO; aqueous sodium hypochlorite in the presence of sodiumbromide and TEMPO (2,2,6,6-tetramethyl-1-piperidinyloxy) free radicalcatalyst; chromium based reagents, such as chromium trioxide, pyridiniumdichromate, or pyridinium chlorochromate; and oxalyl chloride in DMSO inthe presence of a tertiary amine. Examples of reaction-inert solventscomprise dichloromethane, EtOAc, toluene, or pyridine. The oxidation istypically conducted at a temperature of between about −78° C. and about50° C., for between about one and about 24 hours, for example, about 16hours. Such oxidations are well-known to one skilled in the art. See,for example, M. Tamaki, et al., J. Org. Chem., 66, 3593 (2001) and X-I.Qiu, et al., J. Org. Chem., 67, 7162 (2002).

In Scheme 4, Step 3, protected ketocarboxylic acid (IX) is first coupledwith amine (IV), as described hereinabove in Scheme 2, to afford (Va),which is then alkylated to afford ketone (V). The alkylation istypically effected by first forming an enamine by reacting ketone (Va)with a secondary amine, for example, pyrrolidine, piperidine ormorpholine, followed by treatment with an alkylating agent, optionallyin the presence of a base, such as potassium carbonate. Typically, thereaction is effected in a solvent such as benzene, toluene,acetonitrile, or dioxane. Such conversions are well-known to one skilledin the art. See, for example, G. Stork, et al., J. Am. Chem. Soc., 85,207 (1963); M. W. Holladay, et al., J. Med. Chem., 34, 455 (1991); andP. Barraclough, et al., Tetrahedron, 51, 4195 (1995).

In Scheme 4, Step 5, protected ketoester (X), wherein R represents analkyl or arylalkyl moiety, is alkylated under the conditions previouslydescribed in Step 4 to afford ketoester (XI).

In Scheme 4, Step 6, ketoester (XI) is saponified to yield thecorresponding carboxylic acid which, in Step 7, is coupled with anappropriately-substituted amine (IV), as previously describedhereinabove in Scheme 2. The saponification step is typicallyaccomplished by dissolving (XI) in a water-miscible solvent, such asMeOH or EtOH, and water in the presence of a base, such as lithiumhydroxide or sodium hydroxide. The saponification is effected atsuitable temperature, such as between about 0° C. and about 100° C.,preferably room temperature, for a suitable time, such as between aboutone and about 24 hours, for example, about 16 hours.

Preferably, a pharmaceutical composition of the present inventioncomprises a therapeutically effective amount of a compound of Formula(IA), or a pharmaceutically acceptable salt of the compound, or asolvate of the compound or salt, and a pharmaceutically acceptablecarrier, vehicle, diluent or excipient.

More preferably, a pharmaceutical composition of the present inventioncomprises a therapeutically effective amount of the compound(3,3-difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(pyrimidin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)methanone,or a pharmaceutically acceptable salt of said compound, or a solvate ofsaid compound or salt; and a pharmaceutically acceptable carrier,vehicle, diluent or excipient.

The pharmaceutical compositions formed by combining the compounds ofthis invention and the pharmaceutically acceptable carriers, vehicles ordiluents are then readily administered in a variety of dosage forms suchas tablets, powders, lozenges, syrups, injectable solutions and thelike. These pharmaceutical compositions can, if desired, containadditional ingredients such as flavorings, binders, excipients and thelike.

Thus, for purposes of oral administration, tablets containing variousexcipients such as sodium citrate, calcium carbonate and/or calciumphosphate, may be employed along with various disintegrants such asstarch, alginic acid and/or certain complex silicates, together withbinding agents such as polyvinylpyrrolidone, sucrose, gelatin and/oracacia. Additionally, lubricating agents such as magnesium stearate,sodium lauryl sulfate and talc are often useful for tabletting purposes.Solid compositions of a similar type may also be employed as fillers insoft and hard filled gelatin capsules. Preferred materials for thisinclude lactose or milk sugar and high molecular weight polyethyleneglycols. When aqueous suspensions or elixirs are desired for oraladministration, the active pharmaceutical agent therein may be combinedwith various sweetening or flavoring agents, coloring matter or dyesand, if desired, emulsifying or suspending agents, together withdiluents such as water, ethanol, propylene glycol, glycerin and/orcombinations thereof.

For parenteral administration, solutions of the compounds orcompositions of this invention in sesame or peanut oil, aqueouspropylene glycol, or in sterile aqueous solutions may be employed. Suchaqueous solutions should be suitably buffered if necessary and theliquid diluent first rendered isotonic with sufficient saline orglucose. These particular aqueous solutions are especially suitable forintravenous, intramuscular, subcutaneous and intraperitonealadministration. In this connection, the sterile aqueous media employedare all readily available by standard techniques known to those skilledin the art.

For intranasal administration or administration by inhalation, thecompounds or compositions of the invention are conveniently delivered inthe form of a solution or suspension from a pump spray container that issqueezed or pumped by the patient or as an aerosol spray presentationfrom a pressurized container or a nebulizer, with the use of a suitablepropellant, e.g., dichlorodifluoromethane, trichlorofluoromethane,dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In thecase of a pressurized aerosol, the dosage unit may be determined byproviding a valve to deliver a metered amount. The pressurized containeror nebulizer may contain a solution or suspension of a compound of thisinvention. Capsules and cartridges (made, for example, from gelatin) foruse in an inhaler or insufflator may be formulated containing a powdermix of a compound or compounds of the invention and a suitable powderbase such as lactose or starch.

Methods of preparing various pharmaceutical compositions with a certainamount of active ingredient are known, or will be apparent in light ofthis disclosure, to those skilled in this art. For examples of methodsof preparing pharmaceutical compositions, see Remington's PharmaceuticalSciences, Mack Publishing Company, Easton, Pa., 19th Edition (1995).

In another aspect, the invention is directed to a pharmaceuticalcomposition, which comprises a therapeutically effective amount of afirst compound of Formula (I), or a pharmaceutically acceptable salt ofthe compound and a second compound that is an antidiabetic agentselected from insulin and insulin analogs; insulinotropin; biguanides;α₂-antagonists and imidazolines; glitazones; aldose reductaseinhibitors; glycogen phosphorylase inhibitors; sorbitol dehydrogenaseinhibitors; fatty acid oxidation inhibitors; α-glucosidase inhibitors;β-agonists; phosphodiesterase inhibitors; lipid-lowering agents;antiobesity agents; vanadate and vanadium complexes and peroxovanadiumcomplexes; amylin antagonists; glucagon antagonists; growth hormonesecretagogues; gluconeogenesis inhibitors; somatostatin analogs;antilipolytic agents; a prodrug of the antidiabetic agents, or apharmaceutically acceptable salt of the antidiabetic agents and theprodrugs.

In another aspect, the invention is directed to a kit comprising: afirst dosage form comprising a compound of Formula (I), or apharmaceutically acceptable salt of the compound, or a solvate of thecompound or salt; and a second dosage form comprising an antidiabeticagent selected from insulin and insulin analogs; insulinotropin;biguanides; α₂-antagonists and imidazolines; glitazones; aldosereductase inhibitors; glycogen phosphorylase inhibitors; sorbitoldehydrogenase inhibitors; fatty acid oxidation inhibitors; α-glucosidaseinhibitors; β-agonists; phosphodiesterase inhibitors; lipid-loweringagents; antiobesity agents; vanadate and vanadium complexes andperoxovanadium complexes; amylin antagonists; glucagon antagonists;growth hormone secretagogues; gluconeogenesis inhibitors; somatostatinanalogs; antilipolytic agents; prodrugs of the antidiabetic agents, or apharmaceutically acceptable salts of the antidiabetic agents and theprodrug; and a container for containing said first dosage (a) and saidsecond dosage (b). In a preferred embodiment of the kit, both the firstand the second dosage forms independently comprise a pharmaceuticallyacceptable carrier or diluent.

In another aspect, the invention is directed to a therapeutic method ofinhibiting dipeptidyl peptidase-IV comprising administering to a mammalin need of such treatment a therapeutically effective amount of acompound of Formula (I), or a pharmaceutically acceptable salt of thecompound, or a solvate of the compound or salt; either alone or incombination with an antidiabetic agent as described above.

In the present invention, typically, the condition treated is Type 2diabetes, Type 1 diabetes, impaired glucose tolerance, hyperglycemia,metabolic syndrome or a diabetic complication such as diabeticneuropathy, diabetic nephropathy, diabetic retinopathy, diabeticcardiomyopathy and diabetes-related cataracts. Preferably, the conditiontreated is Type 2 diabetes.

In an alternate embodiment, the condition treated is glucosuria,metabolic acidosis, arthritis, obesity, conditions exacerbated byobesity, hypertension, hyperlipidemia, atherosclerosis, osteoporosis,osteopenia, frailty, bone loss, bone fracture, acute coronary syndrome,short stature due to growth hormone deficiency, infertility due topolycystic ovary syndrome, anxiety, depression, insomnia, chronicfatigue, epilepsy, eating disorders, chronic pain, alcohol addiction,diseases associated with intestinal motility, ulcers, irritable bowelsyndrome, inflammatory bowel syndrome short bowel syndrome and cancer.

In another aspect, the invention is directed to a method of identifyingan insulin secretagogue agent for diabetes, comprising: administering anagent of Formula (I) to a fasted, diabetic KK/H1J symptomatic mouse; andassessing a response in the mouse to a subsequent oral glucosechallenge, wherein, if said mouse demonstrates an improvement in thesymptoms, said agent is identified as a treatment for Type 2 diabetes,Type 1 diabetes, impaired glucose tolerance, hyperglycemia, metabolicsyndrome (syndrome X and/or insulin resistance syndrome), glucosuria,metabolic acidosis, arthritis, cataracts, diabetic neuropathy, diabeticnephropathy, diabetic retinopathy, diabetic cardiomyopathy, obesity,conditions exacerbated by obesity, hypertension, hyperlipidemia,atherosclerosis, osteoporosis, osteopenia, frailty, bone loss, bonefracture, acute coronary syndrome, short stature due to growth hormonedeficiency, infertility due to polycystic ovary syndrome, anxiety,depression, insomnia, chronic fatigue, epilepsy, eating disorders,chronic pain, alcohol addiction, diseases associated with intestinalmotility, ulcers, irritable bowel syndrome, inflammatory bowel syndrome;short bowel syndrome, and to prevent disease progression in Type 2diabetes.

The present invention also relates to therapeutic methods for treatingor preventing the above described conditions in a mammal, including ahuman, wherein a compound of Formula (I) of this invention isadministered as part of an appropriate dosage regimen designed to obtainthe benefits of the therapy. The appropriate dosage regimen, the amountof each dose administered and the intervals between doses of thecompound will depend upon the compound of Formula (I) of this inventionbeing used, the type of pharmaceutical compositions being used, thecharacteristics of the subject being treated and the severity of theconditions.

In general, an effective dosage for the compounds of the presentinvention is in the range of 0.01 mg/kg/day to 30 mg/kg/day, preferably0.01 mg/kg/day to 5 mg/kg/day of active compound in single or divideddoses. Some variation in dosage will necessarily occur, however,depending on the condition of the subject being treated. The individualresponsible for dosing will, in any event, determine the appropriatedose for the individual subject. Practitioners will appreciate that “kg”refers to the weight of the patient measured in kilograms.

The compounds or compositions of this invention may be administered insingle (e.g., once daily) or multiple doses or via constant infusion.The compounds of this invention may also be administered alone or incombination with pharmaceutically acceptable carriers, vehicles ordiluents, in either single or multiple doses. Suitable pharmaceuticalcarriers, vehicles and diluents include inert solid diluents or fillers,sterile aqueous solutions and various organic solvents.

The compounds or compositions of the present invention may beadministered to a subject in need of treatment by a variety ofconventional routes of administration, including orally andparenterally, (e.g., intravenously, subcutaneously or intramedullary).Further, the pharmaceutical compositions of this invention may beadministered intranasally, as a suppository, or using a “flash”formulation, i.e., allowing the medication to dissolve in the mouthwithout the need to use water.

EXEMPLIFICATION

Unless noted otherwise, all reactants were obtained commercially.

Flash chromatography was performed according to the method described byW. C. Still et al. in J. Org. Chem. 1978, 43, 2923.

PREPARATIVE EXPERIMENTAL

The compounds and intermediates of the present invention were generallynamed according to the IUPAC (International Union for Pure and AppliedChemistry) recommendations on Nomenclature of Organic Chemistry and theCAS Index rules.

Preparation 1tert-butyl-(2S)-2-[(3,3-Difluoropyrrolidin-1-yl)carbonyl]-4-oxopyrrolidine-1-carboxylateStep1—tert-butyl-(2S,4R)-2-[(3,3-Difluoropyrrolidin-1-yl)carbonyl]-4-hydroxypyrrolidine-1-carboxylate

TEA (0.77 mL, 5.5 mmol) was added to a suspension of3,3-difluoropyrrolidine hydrochloride (0.79 g, 5.5 mmol; Synlett, 55(1995)), in 10 mL of dichloromethane. After five min,(4R)-1-(tert-butoxycarbonyl)-4-hydroxy-L-proline (1.16 g, 5 mmol), HOBt(0.74 g, 5.5 mmol), and EDC (1.05 g, 5.5 mmol) were added. Afterstirring the reaction overnight, the mixture was washed sequentiallywith saturated sodium bicarbonate and brine, dried over magnesiumsulfate, filtered, and concentrated. The residue was purified bychromatography (Biotage® Flash 40S (A Dynax Corp.; Charlottesville,Va.), 9:1 dichloromethane:methanol) to afford 1.07 g of a light pinkfoam. Additional product (0.26 g) was obtained by repeateddichloromethane extractions of the aqueous layer to provide an overallyield of 1.33 g (83%). MS m/z 321 (MH⁺).

Step 2

DMSO (0.57 mL, 8 mmol) in 3 mL dichloromethane was added dropwise to asolution of oxalyl chloride (0.38 mL, 4.4 mmol) in 10 mL dichloromethaneat −65° C. After five min, a solution of the product of Step 1 (1.28 g,4 mmol) in 20 mL dichloromethane was added. After 15 min, TEA (2.79 mL,20 mmol) was added. The reaction mixture was allowed to warm to RT.After 2 hr, the mixture was poured onto ice. The organic layer wasseparated, washed sequentially with 10% NaHCO₃ solution and brine, dried(MgSO₄), and concentrated. The residue was purified by chromatography(Biotage® Flash 40S, 95:5 dichloromethane:MeOH) to afford 765 mg (60%)of the title compound. MS m/z 319 (MH⁺).

Alternatively,tert-butyl-(2S)-2-[(3,3-difluoropyrrolidin-1-yl)carbonyl]-4-oxopyrrolidine-1-carboxylatemay be prepared according to the following procedure.

1-(tert-Butoxycarbonyl)-4-oxo-L-proline (6.88 g, 30 mmol), HOBt (4.46 g,33 mmol), EDC (6.326 g, 33 mmol), and 3,3-difluoropyrrolidinehydrochloride (4.52 g, 31.5 mmol) were dissolved in 100 mL ofdichloromethane and the reaction mixture was cooled to 0° C. in an icebath before adding TEA (8.4 mL, 60 mmol). The reaction mixture was thenallowed to warm to RT. After stirring overnight, saturated sodiumbicarbonate (100 mL) was added and the aqueous layer was extracted withdichloromethane. The combined organic layers were washed with brine,dried over magnesium sulfate, filtered, and concentrated. The residuewas purified by chromatography (Biotage® Flash 40M, eluting with 1:10dichloromethane:hexanes) to afford the title compound 7.85 g (82%yield). MS (El) m/z 319.3 (MH⁺).

Preparation 2tert-Butyl(2S)-2-{[(3R*,4S*)-3,4-difluoropyrrolidin-1-yl]carbonyl}-4-oxopyrrolidine-1-carboxylateStep1—tert-Butyl(2S,4R)-2-{[(3R*,4S*)-3,4difluoropyrrolidin-1-yl]carbonyl}-4-hydroxypyrrolidine-1-carboxylate

(4R)-1-(tert-butoxycarbonyl)-4-hydroxy-L-proline (2.31 g, 10 mmol), wascoupled with (3R,4S)-rel-3,4-difluoropyrrolidine hydrochloride (1.44 g,10 mmol, Preparation 4), in a manner analogous to that described inPreparation 1, Step 1, to afford 2.15 g (67%) of the title product as anoff-white foam. MS m/z 321 (MH⁺).

Step 2

The product of Step 1 (1.97 g, 6.15 mmol) was oxidized in a manneranalogous to that described in Preparation 1, Step 2, to afford 0.74 g(38%) of the title compound as a light yellow solid. MS m/z 319 (MH⁺).

Preparation 3(4S)-1-(tert-Butoxycarbonyl)-4-(4-pyrimidin-2-ylpiperazin-1-yl)-L-proline

1-(tert-Butoxycarbonyl)-4-oxo-L-proline (1.0 g, 4.4 mmol),2-piperazin-1-ylpyrimidine (0.73 g, 4.4 mmol), and acetic acid (275 μL,4.6 mmol) were dissolved in 20 mL of anhydrous 1,2-dichloroethane andsodium triacetoxyborohydride (1.85 g, 8.7 mmol) was added. Afteragitating at RT for 24 hr, the reaction mixture was quenched withsaturated NaHCO₃. The pH was adjusted to pH 7 by addition of solidNaHCO₃ and concentrated HCl, the mixture was extracted withdichloromethane, dried over MgSO₄, filtered, and concentrated to afford1.0 (61 %) of crude material that was sufficiently pure for further use.MS m/z 378 (MH⁺).

Preparation 4 (3R,4S)-rel-3,4-Difluoro-pyrrolidine hydrochloride Step1—2.5-Dihydro-pyrrole-1-carboxylic acid benzyl ester

3-Pyrroline (10 g, 0.145 mol) was added to a slurry of sodiumbicarbonate (14 g, 0.17 mol) in toluene (100 mL). The mixture was cooledto 0° C. and benzyl chloroformate (23 mL, 0.16 mol) was added dropwise.After stirring overnight the solution was diluted with dichloromethane,washed with cold water and brine, dried over magnesium sulfate, andconcentrated to a pale yellow oil that was distilled in vacuo. Bp119-126° C. (0.32 mm).

Step 2—6-Oxa-3-aza-bicyclo[3.1.0]hexane-3-carboxylic acid benzyl ester

The title compound of Step 1 (3.0 g, 15 mmol) was dissolved in a mixtureof acetonitrile (100 mL) and water (70 mL) containing ethylenediaminetetraacetate, disodium salt dihydrate (11 mg, 0.03 mmol). The solutionwas cooled to 0° C. and 1,1,1-trifluoroacetone (14.5 mL, 160 mmol) wasadded over 10 min. Potassium peroxymonosulfate (45 g, 74 mmol) was addedportionwise over 40 min while maintaining the pH at 7 by adding sodiumbicarbonate. The mixture was stirred at 0° C. for 1.5 hr then pouredinto water and extracted with dichloromethane. The combined extractswere dried over magnesium sulfate and concentrated to a colorless oil(3.45 g, 100%).

Step 3—(3RS,4RS)-3-Fluoro-4-hydroxy-pyrrolidine-1-carboxylic acid benzylester

A mixture of TEA trihydrofluoride (1.95 mL, 12 mmol) and the titlecompound of Step 2 (2.62 g, 12 mmol) was heated to 155° C. for three hr,cooled, and partitioned between water and dichloromethane. The aqueousphase was extracted again with dichloromethane and the combined organicextracts were washed with brine, dried over magnesium sulfate andconcentrated. The residue was purified by flash-chromatography (1%methanol in dichloromethane) to give the title compound as a pale oil(1.14 g, 40%).

Step 4—(3R,4S)-rel-3,4-Difluoro-pyrrolidine-1-carboxylic acid benzylester

A solution of the title compound of Step 3 in dichloromethane (15 mL)was cooled to −50° C. and [bis(2-methoxyethyl)amino]sulfur trifluoride(1.3 mL, 6.9 mmol) was added. The solution was warmed to roomtemperature over 18 hr then partitioned between water and EtOAc. Theaqueous phase was extracted again with EtOAc and the combined organicextracts were washed with brine, dried over magnesium sulfate, andconcentrated. The residue was purified by flash-chromatography(dichloromethane) to give the product as a brown oil (1.14 g, 40%).

Step 5

A solution of the title compound of Step 4 (675 mg, 2.8 mmol) in EtOH(10 mL) containing 10% Pd/C (200 mg) was hydrogenated at 40 psi in aParr apparatus for 18 hr. The solution was filtered over diatomaceousearth and the filtrate was concentrated to dryness, leaving a yellowsolid (400 mg, 100%).

Preparation 5(S)-2-(3-Fluoro-azetidine-1-carbonyl)-4-oxo-pyrrolidine-1-carboxylicacid tert-butyl ester Step 1—Benzhydryl-3-fluoro-azetidine hydrochloride

1-Benzyhydryl-azetidin-3-ol (5.0 g, 20.9 mmol) was dissolved in 50 mL ofbenzene, the solution cooled to 15° C., and (diethylamino)sulfurtrifluoride (10.1 g, 62.7 mmol) was added dropwise. After stirringovernight at room temperature, saturated sodium bicarbonate was added.The mixture was extracted with EtOAc, dried over magnesium sulfate,filtered, and concentrated. The residue was purified by chromatography(Biotage® 40S,10% EtOAc/hexanes). The product was dissolved in EtOAc,treated with HCl (15 mL, 2N in ether), heated briefly, and concentrated.The solid was triturated with ether, filtered, and dried to provide 2.58g of the title compound. MS m/z 242.3 (MH⁺).

Step 2—3-Fluoro-azetidine hydrochloride

A solution of the product of Step 1 (2.58 g, 9.3 mmol) in 30 mL ofmethanol containing 10% Pd/C (0.38 g) was hydrogenated at 30-50 psi in aParr apparatus for 60 hr. The solution was filtered over diatomaceousearth and the filtrate concentrated to dryness. The solid wasrecrystallized from MeOH/EtOAc to furnish 0.62 g (60%) of the titlecompound.

Step 3

N-tert-Boc-4-oxo-L-proline (917 mg, 4 mmol), the title compound of Step2 (446 mg, 4 mmol), and HATU (1.673 g, 4.4 mmol) were mixed undernitrogen in anhydrous methylene chloride. The solution was cooled in anice bath before the addition of DIEA (1.4 mL, 8 mmol). The reactionmixture was allowed to warm to RT and stirred overnight. Saturatedsodium bicarbonate was added, the phases were separated and the aqueousphase was extracted with methylene chloride. The combined organicportions were washed with brine and dried over magnesium sulfate. Thecrude product (2.11 g) was purified by chromatography (Biotage® Flash40S, 95:5 EtOAc:MeOH) to give the title product as light pink foam (1.06g, 92%). MS m/z 287.3 (MH⁺).

Preparation 6(S)-2-((S)-3-Fluoro-pyrrolidine-1-carbonyl)-4-oxo-pyrrolidine-1-carboxylicacid tert-butyl ester

N-tert-Boc-4-oxo-L-proline (2.29 g, 10 mmol), (S)-3-fluoropyrrolidinehydrochloride (1.38 g, 11 mmol) and TEA (2.09 mL, 15 mmol) were mixed inanhydrous methylene chloride (30 mL) under nitrogen. HOBT (2.03 g, 15mmol) was added and the mixture cooled to 0° C. in an ice bath beforeaddition of EDC (2.10, 11 mmol). The reaction mixture was allowed towarm to RT and stirred overnight. The mixture was washed with saturatedsodium bicarbonate and brine and dried over magnesium sulfate. The crudematerial (3.15 g) was recrystallized from hexane:EtOAc (2:1) to give thetitle compound as light yellow needles (2.18 g, 73%). MS m/z 301.3(MH⁺).

Preparation 7(2S,4S)-2-(3,3-Difluoro-pyrrolidine-1-carbonyl)-4-piperazin-1-yl-pyrrolidine-1-carboxylicacid tert-butyl ester Step1—4-[(3S,5S)-1-tert-Butoxycarbonyl-5-(3,3-difluoro-pyrrolidine-1-carbonyl)-pyrrolidin-3-yl]-piperazine-1-carboxylicacid benzyl ester

To a solution of the title compound of Preparation 1 (1.59 g, 5 mmol)and 1-(benzyloxycarbonyl)piperazine (1.21 g, 5.5 mmol) in1,2-dichloroethane (20 mL) was added AcOH (0.3 mL, 1.05 equiv.),followed by sodium triacetoxyborohydride (2.119 g, 10 mmol). Thereaction mixture was stirred at RT for 4 hr. Saturated sodiumbicarbonate was added and the product extracted with methylene chloride.The organic phase was washed with brine and dried over magnesiumsulfate. After evaporation, the crude product (2.28 g yellow foam) waspurified by flash chromatography eluting with EtOAc to give titlecompound as white foam (1.28 g, 49%). MS m/z 523.3 (MH⁺).

Step 2

The product of Step 1 (1 g, 1.91 mmol) was dissolved in EtOH (50 mL) and10% Pd/C (1 g, 1 equiv. w/w) was carefully added, followed by1,4-cyclohexadiene (1.81 mL, 10 equiv.). The mixture was stirred gentlyin a tightly-capped flask at RT overnight. The reaction mixture wasfiltered through diatomaceous earth and concentrated to give the productas yellow semisolid (758 mg, 100%). MS m/z 389.4 (MH⁺).

Preparation 8(S)-2-(3,3-Difluoro-azetidine-1-carbonyl)-4-oxo-pyrrolidine-1-carboxylicacid tert-butyl ester

N-tert-BOC4-oxo-L-proline (458 mg, 2 mmol), 3,3-difluoroazetidinehydrochloride (258 mg, 2 mmol)(prepared as described in WO 2000/47582),and DIPEA (0.35 mL, 2 mmol) were mixed in anhydrous methylene chloride(10 mL) and cooled to 0° C. HOBT (405 mg, 3 mmol) was then added in oneportion followed by EDC hydrochloride (422 mg, 2.2 mmol). The resultingmixture was allowed to warm to RT and stirred overnight. Saturatedsodium bicarbonate was added, the organic layer was separated, and theaqueous phase extracted with methylene chloride. The combined organicextracts were washed twice with brine, dried over magnesium sulfate,filtered, and concentrated. The crude product (570 mg) was trituratedwith hexanes:methylene chloride (10:1), filtered, and dried in a vacuumoven to afford 510 mg (84% yield) of the title product as a light orangepowder. MS (m/z): 305.1 (MH⁺).

Preparation 9 (2S,4S)-4-Fluoro-pyrrolidine-2-carbonitrile hydrochlorideStep 1—(2S,4S)-4-Fluoro-pyrrolidine-1,2-dicarboxylic acid 2-tert-butylester 1-(2,5-dioxo-pyrrolidin-1-yl)ester

To a solution of N-tert-BOC-cis-4-fluoro-L-proline (700 mg, 3 mmol) inanhydrous DMF (8 mL) was at 0° C. added N-hydroxysuccinimide (380 mg,3.3 mmol) in one portion, followed by 1,3-diisopropylcarbodiimide (391mg, 3.1 mmol) in small portions. The reaction was allowed to warm to RTand stirred overnight. The mixture was diluted with 100 mL of water, theprecipitate was collected, washed with cold water, and dried in a vacuumoven overnight. The product (1.093 g) was used without furtherpurification. MS m/z 331.3 (MH⁺).

Step 2—(2S,4S)-2-Carbamoyl-4-fluoro-pyrrolidine-1-carboxylic acidtert-butyl ester

The title compound of Step 1 (1.03 g, 3.12 mmol) was dissolved indioxane (12 mL) at RT and the solution was treated with concentratedaqueous ammonium hydroxide (10 mmol) dropwise. The resulting thicksolution was stirred at RT for three hr, then acidified with 6N HCl topH 4-5, and extracted with methylene chloride (2×). The combinedextracts were washed with saturated sodium bicarbonate and brine, driedover magnesium sulfate, filtered, and concentrated to afford 562 mg (78%yield) of a clear oil. MS m/z 233.3 (MH⁺).

Step 3—(2S,4S)-2-Cyano-4-fluoro-pyrrolidine-1-carboxylic acid tert-butylester

To a solution of the title compound of Step 2 (550 mg, 2.37 mmol) anddry pyridine (0.4 mL, 2 equiv.) in anhydrous methylene chloride (15 mL)at 0° C. was added a solution of TFAA in 2 mL of methylene chlorideunder nitrogen. The solution was stirred at 0° C. for two hr and then atRT for one hr. The reaction mixture was washed with saturated aqueoussodium bicarbonate and brine, dried over magnesium sulfate, filtered,and concentrated. The residue was purified by flash chromatography onsilica gel to give 458 mg (90% yield) of an oil that solidified onstanding. MS m/z 215.3 (MH⁺).

Step 4

The title compound of Step 3 (400 mg) was dissolved in dry acetonitrile(8 mL) and 0.5 mL of 4N HCl in dioxane was added under nitrogen. Theresulting solution was stirred at RT overnight and the white precipitatethat formed was filtered and dried in a vacuum oven to yield 128 mg (46%yield) of the title compound. MS m/z 115.1 (MH⁺). Additional productcould be obtained from the filtrate.

Preparation 10 (2S)-4,4-Difluoro-pyrrolidine-2-carbonitrilehydrochloride Step 1—N-tert-BOC-4,4-Difluoropyrrolidine-2-carbonitrile

To a solution of N-tert-BOC4,4-difluoropyrrolidine-L-proline amide (250mg, 1 mmol) and dry pyridine (97 μL, 1.2 equiv.) in anhydrous methylenechloride at 0° C. was added a solution of TFAA (252 mg, 1.2 equiv.) in 1mL of anhydrous methylene chloride. The solution was allowed to warm toRT and stirred for 36 hr. The reaction was quenched with saturatedammonium chloride, the organic phase was washed successively with 1NHCl, saturated sodium bicarbonate and brine, dried over magnesiumsulfate, filtered, and concentrated to afford 252 mg of a whitesemisolid. MS m/z 233.1 (MH⁺).

Step 2

The title compound of Step 1 (245 mg) was dissolved in dry acetonitrile(10 mL) and 0.5 mL of 4N HCl was added. The resulting solution wasstirred at RT for five hr and the solvents were removed. The residue wastriturated with EtOAc, the solid was filtered, and then dried underhigh-vacuum to afford 105 (59% yield) of the title compound as a whitesolid. MS m/z 133.2 (MH⁺).

The compounds of formula (I), the stereoisomers thereof, and thepharmaceutically acceptable salts of the compounds and stereoisomers,may be prepared as described in the following Examples. The free basecompounds of the present invention may be obtained from their salt formsby conventional means such as disclosed in Example 113, herein.

Example 1((2S,4S)-4-(4-(3-(Trifluoromethyl)phenyl)piperazin-1-yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanonedihydrochloride Step1—tert-Butyl(2S,4S)-2-[(3,3-difluoropyrrolidin-1-yl)carbonyl]-4-{4-[3-(trifluoromethyl)phenyl]piperazin-1-yl}pyrrolidine-1-carboxylate

The title compound of Preparation 1 (96 mg, 0.3 mmol),1-[3-(trifluoromethyl)phenyl]piperazine (70 mg, 0.3 mmol) and AcOH (18μL, 0.3 mmol) were dissolved in 8 mL anhydrous 1,2-dichloroethane.Sodium triacetoxyborohydride (127 mg, 0.6 mmol) was added. Afterstirring the reaction at RT for 3 hr, the reaction was quenched withsaturated sodium bicarbonate, extracted with EtOAc, washed with brine,dried over magnesium sulfate, filtered, and concentrated. The crudematerial was purified by chromatography (Biotage® Flash 40S, 95:5dichloromethane:MeOH) to afford 126 mg (79%) of the title compound as awhite foam. MS m/z 533 (MH⁺).

Step 2

The product of Step 1 (120 mg, 0.225 mmol) was treated with 4N HCl indioxane (5 mL). After two hr at RT, the mixture was concentrated todryness, triturated with ether, filtered, and dried in vacuo to provide92 mg of the title compound as a white solid. MS m/z 433 (MH⁺).

Using appropriate starting materials, the hydrochloride salts of thecompounds of Examples 2 to 112, disclosed in Table 1 hereinbelow, wereprepared in a manner analogous to that described in Example 1. TABLE 1Example Name MS(M + 1) 2((2S,4S)-4-(4-(5-(Trifluoromethyl)pyridin-2-yl)piperazin-1- 434yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 3((2S,4S)-4-(4-(5-(Trifluoromethyl)pyridin-2-yl)-1,4-diazepan-1- 448yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 4((2S,4S)-4-(4-(3-(Trifluoromethyl)phenyl)piperazin-1-yl)pyrrolidin-2-433 yl)-((3R*,4S*)-3,4-difluoropyrrolidin-1-yl)-methanone 5((2S,4S)-4-(4-(2-(Trifluoromethyl)quinolin-4-yl)piperazin-1- 484yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 6(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(5-nitropyridin-2- 411yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 7((2S,4S)-4-(4-(3-Cyanopyridin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)- 391(3,3-difluoropyrrolidin-1-yl)-methanone 8((2S,4S)-4-(4-(5-(Trifluoromethyl)pyridin-2-yl)piperazin-1- 434yl)pyrrolidin-2-yl)-((3R*,4S*)-3,4-difluoropyrrolidin-1-yl)-methanone 9((2S,4S)-4-(4-(3-Cyanopyridin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)- 391((3R*,4S*)-3,4-difluoropyrrolidin-1-yl)-methanone 10((2S,4S)-4-(4-(3-Cyanopyrazin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)- 392((3R*,4S*)-3,4-difluoropyrrolidin-1-yl)-methanone 11((2S,4S)-4-(4-(4-(Trifluoromethyl)phenyl)piperazin-1-yl)pyrrolidin-2-433 yl)-((3R*,4S*)-3,4-difluoropyrrolidin-1-yl)-methanone 12((2S,4S)-4-(2-(Trifluoromethyl)-5,6-dihydroimidazo[1,2-a]pyrazin- 3947(8H)-yl)pyrrolidin-2-yl)-((3R*,4S*)-3,4-difluoropyrrolidin-1-yl)-methanone 13((2S,4S)-4-(4-(3-Cyanopyrazin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)- 392(3,3-difluoropyrrolidin-1-yl)-methanone 14((2S,4S)-4-(2-(Trifluoromethyl)-5,6-dihydroimidazo[1,2-a]pyrazin- 3947(8H)-yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 15((3R*,4S*)-3,4-difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(pyrimidin-2- 367yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 16((2S,4S)-4-(4-(2-(Trifluoromethyl)phenyl)piperazin-1-yl)pyrrolidin-2-433 yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 17((2S,4S)-4-((1S,5R,6R)-6-Amino-3-aza-bicyclo[3.1.0]hexan-3- 301yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 18((2S,4S)-4-(4-Cyano-4-phenylpiperidin-1-yl)pyrrolidin-2-yl)-(3,3- 389difluoropyrrolidin-1-yl)methanone 19((2S,4S)-4-(4-(1,1-Dioxo-1H-1,2-benzo[d]isothiazol-3-yl)-piperazin- 4541-yl)pyrrolidin-2-yl)-(3,3-difluoro-pyrrolidin-1-yl)-methanone 20((2S,4S)-4-(4-(5-(Trifluoromethyl)-1,3,4-thiadiazol-2-yl)piperazin-1-441 yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 21(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(isothiazol-3-yl)piperazin-372 1-yl)pyrrolidin-2-yl)methanone 22(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(8-methyl- 421[1,2,4]triazolo[4,3-a]pyrazin-3-yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 23((2S,4S)-4-(3-(Trifluoromethyl)-5,6-dihydro-[1,2,4]triazolo[4,3- 395a]pyrazin-7(8H)-yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 24(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(2,6-dimethylpyrimidin-4-395 yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 25((2S,4S)-4-(4-(Benzo[d]isothiazol-3-yl)piperazin-1-yl)pyrrolidin-2-yl)-422 (3,3-difluoropyrrolidin-1-yl)-methanone 26((2S,4S)-4-(4-(4-(Trifluoromethyl)-6-methylpyridin-2-yl)piperazin-1- 448yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 27(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(oxazolo[5,4-b]pyridin-2-407 yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 28(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(4-methylpyrimidin-2- 381yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 29((2S,4S)-4-(4-(4-Cyanopyridin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)- 391(3,3-difluoropyrrolidin-1-yl)-methanone 30((2S,4S)-4-(4-(7-(Trifluoromethyl)quinolin-4-yl)piperazin-1- 484yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 31((2S,4S)-4-(4-(5-Cyanopyridin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)- 391(3,3-difluoropyrrolidin-1-yl)-methanone 32(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(pyridin-2-yl)piperazin-1-366 yl)pyrrolidin-2-yl)methanone 33((2S,4S)-4-(4-(6-(Trifluoromethyl)quinolin-4-yl)piperazin-1- 484yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 34(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(5-methylpyridin-2- 380yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 35((2S,4S)-4-(4-(4-(Trifluoromethyl)pyrimidin-2-yl)piperazin-1- 435yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 36(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(3-methyl-1,2,4-oxadiazol-370 5-yl)piperidin-1-yl)pyrrolidin-2-yl)-methanone 37(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(quinolin-2-yl)piperazin-1-416 yl)pyrrolidin-2-yl)-methanone 38(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(6-methoxypyridin-2- 396yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 39(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(5-methyl-1,2,4-oxadiazol-370 3-yl)piperidin-1-yl)pyrrolidin-2-yl)-methanone 40(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(quinolin-8-yl)piperazin-1-416 yl)pyrrolidin-2-yl)-methanone 41(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(1-phenyl-1H-imidazol-2- 430yl)piperidin-1-yl)pyrrolidin-2-yl)-methanone 42(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(quinoxalin-5-yl)piperazin-417 1-yl)pyrrolidin-2-yl)-methanone 43((2S,4S)-4-(4-(Benzo[d]isoxazol-3-yl)piperazin-1-yl)pyrrolidin-2-yl)-406 (3,3-difluoropyrrolidin-1-yl)methanone 44(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(8-trifluoromethyl-3,4- 444dihydro-1H-benzo[4,5]imidazo[1,2-a]pyrazin-2-yl)-pyrrolidin-2-yl]-methanone 45(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-phenylpiperidin-1- 364yl)pyrrolidin-2-yl)-methanone 46((2S,4S)-4-(4-(3-(Trifluoromethyl)phenyl)piperidin-1-yl)pyrrolidin-2-432 yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 47((2S,4S)-4-(4-(3-(Trifluoromethyl)pyridin-2-yl)piperazin-1- 434yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 48((2S,4S)-4-(4-(4-(Trifluoromethyl)quinolin-2-yl)piperazin-1- 484yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 49((2S,4S)-4-(2-(Trifluoromethyl)-7,8-dihydropyrido[4,3-d]pyrimidin- 4066(5H)-yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 50(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(4-methyl-6- 457phenylpyrimidin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 51((2S,4S)-4-(4-(1H-Benzo[d][1,2,3]triazol-1-yl)piperidin-1- 405yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone 52(3,3-Difluoropyrrolidin-1-yl)((2S,4S)-4-(4-(thiazol-2-yl)piperazin-1-372 yl)pyrrolidin-2-yl)-methanone 53(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(3-methylpyridin-2- 380yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 54((2S,4S)-4-(4-(Benzo[d]oxazol-2-yl)piperazin-1-yl)pyrrolidin-2-yl)- 406(3,3-difluoropyrrolidin-1-yl)-methanone 55(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(6-phenylpyridin-2- 442yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 56(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-((3R,5S)-3,5-dimethyl-4- 424(4,6-dimethyl-1,3,5-triazin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 57[(2S,4S)-4-(2-Cyclopropyl-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6- 378.4yl)-pyrrolidin-2-yl]-(3,3-difluoro-pyrrolidin-1-yl)-methanone 58(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-methoxy-7,8-dihydro-5H-368.3 pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone 59(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-phenyl-7,8-dihydro-5H-414.4 pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone 60(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[4,5-c]pyridin-2-407.4 yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone 61(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-c]pyridin-2-407.4 yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone 62(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2,3,4,5-tetrahydro- 367.4[1,2′]-bipyrazinyl-4-yl)-pyrrolidin-2-yl]-methanone 63{(2S,4S)-4-[4-(3,5-Dichloro-pyridin-4-yl)-piperazin-1-yl]-pyrrolidin-2-434.2 yl}-(3,3-difluoro-pyrrolidin-1-yl)-methanone 64(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-quinoxalin-2-yl-piperazin-417.4 1-yl)-pyrrolidin-2-yl]-methanone 654-[(3S,5S)-5-(3,3-Difluoro-pyrrolidine-1-carbonyl)-pyrrolidin-3-yl]-396.3 piperazine-1-sulfonic acid dimethylamide 66[(2S,4S)-4-(2-Amino-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6-yl)- 353.3pyrrolidin-2-yl]-(3,3-difluoro-pyrrolidin-1-yl)-methanone 67(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-methyl-4-pyrimidin-2-yl-381.4 piperazin-1-yl)-pyrrolidin-2-yl]-methanone 68(3,3-Difluoro-pyrrolidin-1-yl)-{(2S,4S)-4-[4-(5-ethyl-pyrimidin-2-yl)-395.4 piperazin-1-yl]-pyrrolidin-2-yl}-methanone 69{(2S,4S)-4-[4-(5-Bromo-pyrimidin-2-yl)-piperazin-1-yl]-pyrrolidin-2-445.4 yl}-(3,3-difluoro-pyrrolidin-1-yl)-methanone 704-[(3S,5S)-5-(3,3-Difluoro-pyrrolidine-1-carbonyl)-pyrrolidin-3-yl]-423.4 piperazine-1-carboxylic acid benzyl ester 71((2S,4S)-4-(2-(4-Chlorophenyl)-7,8-dihydropyrido[4,3-d]pyrimidin- 448.46(5H)-yl)pyrrolidin-2-yl)(3,3-difluoropyrrolidin-1-yl)methanone 72(3,3-Difluoropyrrolidin-1-yl)((2S,4S)-4-(7,8-dihydro-2- 382.4propylpyrido[4,3-d]pyrimidin-6(5H)-yl)pyrrolidin-2-yl)methanone 73{(2S,4S)-4-[4-(5-Chloro-benzooxazol-2-yl)-piperazin-1-yl]-pyrrolidin-440.4 2-yl}-(3,3-difluoro-pyrrolidin-1-yl)-methanone 74(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-((2-pyridin-2-yl)-7,8-dihydro-415.4 5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone 75(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-pyridin-4-yl-7,8-dihydro-415.4 5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone 76(3,3-Difluoro-pyrrolidin-1-yl)-{(2S,4S)-4-[4-(5-methyl-benzooxazol-2-420.4 yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone 77{(2S,4S)-4-[4-(6-Chloro-benzooxazol-2-yl)-piperazin-1-yl]-pyrrolidin-440.4 2-yl}-(3,3-difluoro-pyrrolidin-1-yl)-methanone 78(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(7,8-dihydro-5H-pyrido[4,3-338.4 d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone 79((S)-3-Fluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-c]pyridin-2-389.4 yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone 804-[(3S,5S)-5-((S)-3-Fluoro-pyrrolidine-1-carbonyl)-pyrrolidin-3-yl]-374.4 3,4,5,6-tetrahydro-2H-[1,2′]bipyrazinyl-3′-carbonitrile 81((S)-3-Fluoro-pyrrolidin-1-yl)-{(2S,4S)-4-[4-(5-trifluoromethyl-pyridin-416.4 2-yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone 82((S)-3-Fluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-b]pyridin-2-389.4 yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone 832-{4-[(3S,5S)-5-((S)-3-Fluoro-pyrrolidine-1-carbonyl)-pyrrolidin-3-yl]-373.4 piperazin-1-yl}-nicotinonitrile 84((S)-3-Fluoro-pyrrolidin-1-yl)-{(2S,4S)-4-[4-(3-trifluoromethyl-pyridin-416.5 2-yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone 85((2S,4S)-4-(2-(Trifluoromethyl)-7,8-dihydropyrido[4,3-d]pyrimidin- 388.46(5H)-yl)pyrrolidin-2-yl)((S)-3-fluoropyrrolidin-1-yl)methanone 86((S)-3-Fluoro-pyrrolidin-1-yl)-{(2S,4S)-4-[4-(4-methyl-pyrimidin-2-yl)-363.5 piperazin-1-yl]-pyrrolidin-2-yl}-methanone 87((S)-3-Fluoropyrrolidin-1-yl)((2S,4S)-4-(4-(pyrazin-2-yl)piperazin-1-349.4 yl)pyrrolidin-2-yl)methanone 88[(2S,4S)-4-(2-Cyclopropyl-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6- 360.4yl)-pyrrolidin-2-yl]-((S)-3-fluoro-pyrrolidin-1-yl)-methanone 89((S)-3-Fluoro-pyrrolidin-1-yl)-{(2S,4S)-4-[4-(2-trifluoromethyl- 466.5quinolin-4-yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone 90(3-Fluoroazetidin-1-yl)((2S,4S)-4-(4-(pyrazin-2-yl)piperazin-1- 335.4yl)pyrrolidin-2-yl)methanone 914-[(3S,5S)-5-(3-Fluoro-azetidine-1-carbonyl)-pyrrolidin-3-yl]-3,4,5,6-360.4 tetrahydro-2H-[1,2′]bipyrazin-3′-carbonitrile 92(3-Fluoro-azetidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-b]pyridin-2-yl- 375.4piperazin-1-yl)-pyrrolidin-2-yl]-methanone 93(3-Fluoro-azetidin-1-yl)-{(2S,4S)-4-[4-(3-trifluoromethyl-pyridin-2-yl)-402.4 piperazin-1-yl]-pyrrolidin-2-yl}-methanone 94(3-Fluoro-azetidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-c]pyridin-2-yl- 375.4piperazin-1-yl)-pyrrolidin-2-yl]-methanone 95[(2S,4S)-4-(2-Cyclopropyl-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6- 346.4yl)-pyrrolidin-2-yl]-(3-fluoro-azetidin-1-yl)-methanone 962-{4-[(3S,5S)-5-(3-Fluoro-azetidine-1-carbonyl)-pyrrolidin-3-yl]- 359.4piperazin-1-yl}-nicotinonitrile 97(3-Fluoroazetidin-1-yl)((2S,4S)-4-(4-(5-(trifluoromethyl)pyridin-2-402.4 yl)piperazin-1-yl)pyrrolidin-2-yl)methanone 98(3-Fluoro-azetidin-1-yl)-[(2S,4S)-4-(2-trifluoromethyl-7,8-dihydro-374.4 5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone 99(3-Fluoro-azetidin-1-yl)-{(2S,4S)-4-[4-(4-methyl-pyrimidin-2-yl)- 349.4piperazin-1-yl]-pyrrolidin-2-yl}-methanone 100(3-Fluoro-azetidin-1-yl)-{(2S,4S)-4-[4-(2-trifluoromethyl-quinolin-4-452.5 yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone 101[(2S,4S)-4-(4-Benzooxazolo-2-ylpiperazin-1-yl)-pyrrolidin-2-yl]- 406.4((3R*,4S*)-3,4-difluoro-pyrrolidin-1-yl)-methanone 102((3R*,4S*)-3,4-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-trifluoromethyl-406.4 7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone 103((3R*,4S*)-3,4-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-407.4 c]pyridin-2-yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone 104((3R*,4S*)-3,4-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-407.4 b]pyridin-2-yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone 105((3R*,4S*)-3,4-Difluoro-pyrrolidin-1-yl)-{(2S,4S)-4-[4-(4-methyl- 381.4pyrimidin-2-yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone 106(3,3-Difluoro-azetidin-1-yl)-{(2S,4S)-4-[4-(3-trifluoromethyl-pyridin-420.2 2-yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone 1072-{4-[(3S,5S)-5-(3,3-Difluoro-azetidin-1-carbonyl)-pyrrolidin-3-yl]-377.2 piperazin-1-yl}-nicotinonitrile 108(3,3-Difluoro-azetidin-1-yl)-[(2S,4S)-4-(2-trifluoromethyl-7,8- 392.2dihydro5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone 109(3,3-Difluoro-azetidin-1-yl)-{(2S,4S)-4-[4-(2-trifluoromethyl-quinolin-470.2 4-yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone 110(3,3-Difluoro-azetidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-c]pyridin-2-yl-393.2 piperazin-1-yl)-pyrrolidin-2-yl]-methanone 111{(2S,4S)-4-[5-(4-Chloro-phenyl)-2-aza-bicyclo[2.2.1]hept-2-yl]- 410.2pyrrolidin-2-yl}-(3,3-difluoro-pyrrolidin-1-yl)-methanone 112(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-trifluoromethyl-5,8- 406.1dihydro-6H-pyrido[3,4-d]pyrimidin-7-yl)-pyrrolidin-2-yl]-methanone

Example 113(3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(pyrimidin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone

Step1—(S)-2-(3,3-Difluoro-pyrrolidine-1-carbonyl)-4-oxo-pyrrolidine-1-carboxylicacid tert-butyl ester

(S)-4-Oxo-pyrrolidine-1,2-dicarboxylic acid 1-tert-butyl ester (6.6 kg,1.0 equivalent) was charged to a reactor, followed by addition ofdichloromethane (15 volumes). The reaction mixture was cooled to 0° C.Triethylamine (4.82 liters, 1.2 equiv) was added over 30 minutes. Themixture turned from suspension to a clear solution at the end oftriethylamine addition. The mixture was held at 0° C. to 5° C. for 10minutes. Pivaloyl chloride (3.65 kg, 1.05 equivalents) was added slowlywhile keeping the reaction temperature at 0° C. to 5° C. The reactionmixture turned back to aslurry. The reaction mixture was sampled forcompletion by HPLC (using diethylamine to derivatize) after held for 1hour at 0° C. to 5° C. 3,3-Difluoro-pyrrolidine hydrochloride (4.13 kg,1.0 equivalent) was charged to the above mixture over 10 minutes at −10°C. to 0° C. Triethylamine (4.0 liters, 1.0 equiv) was introduced slowlyover 70 minutes at −10° C. to 0° C. Upon completion of triethylamineaddition, the mixture was stirred for 1 h at 0 to 5° C. The reaction wascomplete by HPLC assay (˜1% starting material). The reaction wasquenched with water (10 volumes) at 0° C. to 5° C. The mixture washeated to 20° C. to 25° C. The layers were separated, and the organiclayer was washed with 0.5 M HCl (5 volumes). The organic layer was againwashed with combined 5% NaHCO₃ (2 volumes) and half saturated brinesolution (1.64 M, 3 volumes). The organic solution was concentratedatmospherically to a low stirrable volume (approximately 20 liters).Ethyl acetate (12.6 volumes, 82.8 liters) was added, the solution wasconcentrated atmospherically to ˜6 volumes. The mixture was held at 60°C. to 65 ° C. for 2 hours and cooled to room temperature over 3 hours.The mixture was held at 20° C. to 25° C. for 8 hours. Heptane (8volumes) was added, and the mixture was granulated for a minimum of 2hours. The solid was filtered, rinsed with 2:1 heptane/ethyl acetate (1volume), and dried in a tray dryer at 25° C. to 35° C. for a minimum of12 h. Yield: 7.26 kg, 79%. HPLC purity: 99.7%. The mother liquor (86liters) was concentrated to 12 liters under partial vacuum at 65° C. to70° C. The mixture was cooled to 60° C. to 65° C. Ethyl acetate (4.0liters) was added slowly over 15 minutes. The mixture was cooled to 20°C. to 25° C. over 2 hours and was held at that temperature for at least2 hours. The solid was filtered and rinsed with heptane/ethyl acetate(3:1 v/v, 1.7 liters). Drying in a tray dryer for 12 hours at 35° C. to45 ° C. yielded 435 grams of product. HPLC purity: 96.4%.

Step2—(2S,4S)-2-(3,3-Difluoro-pyrrolidine-1-carbonyl)-4-(4-pyrimidin-2-yl-piperazin-1-yl)-pyrrolidine-1-carboxylicacid tert-butyl ester

A reactor was charged with THF (20 volumes), 2-piperazin-1-yl-pyrimidine(2.17 kg, 1.05 equivalents) and the product from Step 1 (4.00 kg, 1.0equivalent). The mixture was held at 20° C. to 25° C. until all materialwas dissolved over 30 minutes. Acetic acid (0.792 kg, 1.05 equivalents)as added. The mixture was stirred for 1 hour during which the reactionmixture turned to cloudy. The reaction mixture was refluxed for 30minutes and then concentrated at 60° C. to 70° C. until a steadytemperature of 66.9° C. was observed in the overheads indicatingcomplete removal of water from the system. More THF was added asnecessary. At the end, THF was added to bring the total volume in thereactor to 15 volumes of the limit reagent. The reaction mixture wascooled to −3° C. to 7° C. and sampled for complete formation of imine byHPLC (using sodium triacetoxyborohydride to reduce imine). Sodiumtriacetoxyborohydride (5.33 kg, 2.0 equivalents) was added portion-wiseto the suspension at −5° C. to 15° C. The reaction mixture was heated to20° C. to 25° C. and held for 12 hours. HPLC results confirmed thereaction was complete by 99.8%. Sodium bicarbonate aqueous solution (10%w/w, 10 volumes) was added. The slurry was concentrated to remove 10volumes of THF under partial vacuum at 30° C. to 60° C. Ethyl acetate(10 volumes) was added to the suspension after it cooled to 20° C. to25° C. The organic phase was separated and the aqueous phase was checkedby HPLC. It contained less than 2% of the product. The organic phase waswashed with water (5 volumes), saturated brine solution (5 volumes) andconcentrated to a small volume (2 volumes) under partial vacuum at 45°C. to 50° C. To the slurry was added heptane (10 volumes) at 45° C. to50° C. over 30 minutes. The mixture was cooled to 20° C. to 25° C. andgranulated for 2 hours. Solid was collected by filtration, rinsed withheptane (2 volumes). Drying in a tray dryer for 12 hours at 35° C. to45° C. yield 5.35 kg (91.3%) of the product.

Step3—(3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-pyrimidin-2-yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone

Water (19 liters, 2 volumes) was charged to a reactor followed by theproduct from Step 2 (9.57 kg, 1.0 equivalent). To the slurry was addedconcentrated HCl (37 wt % in water, 19.1 liters, 2 volumes) slowly at20° C. to 30° C. over 4 hours. The slurry went into solution after 12liters of HCl was added. After the addition completion, the reaction wascomplete by HPLC assay. The reaction mixture was cooled to 5° C. to 15°C. To the mixture was added 50% NaOH aqueous solution slowly withagitation to pH 10 to pH 11. The pH was monitored with a pH meterclosely during the neutralization. The total volume of 50% NaOH addedwas 12.45 liters. The mixture was warmed to 20° C. to 25° C. andextracted with ethyl acetate twice (115 liters, 12 volumes and 57liters, 6 volumes, respectively). The sample from aqueous layer aftersecond extraction was analyzed by HPLC and showed only 1% of the productin that aqueous solution. The organic layers were combined and treatedwith magnesium sulfate (5 kg) for 1 hour. The mixture was filtered. Thefilter cake was rinsed with ethyl acetate (10 liters). The filtrate wascharged back to the reactor via a 0.2 micron in-line filter for speckfree operation. (The following operations were performed under speckfree conditions.) The solution was concentrated to 20 liters (2 volumes)under partial vacuum at 50° C. to 60° C. The mixture was cooled to 20°C. to 25° C. over 30 minutes. Upon cooling to room temperature,crystallization occurred. The mixture was held for 30 minutes. Hexanes(20 liters, 2 volumes) was added slowly over 1 hour. The mixture wasgranulated for 2 hours. The solid product was collected by filtrationand rinsed with hexanes/ethyl acetate (10 liters, 1:1 v/v). The filterwas blown dry with nitrogen for a minimum of 2 hours. The product wasdried in a tray dryer at 44° C. for 12 hours. Yield: 5.7 kg, 75.9%. m.p.156° C. MS m/z 367 (MH⁺). ¹H NMR (400 MHz, D₂O): δ 8.15 (d, 2H, J=5.0Hz, CH of pyrimidine), 6.55 (t, 1H, J=4.8 Hz, CH ofpyrimidine),3.87-3.81 (dd, 1H, H_(2b) of proline, rotomeric), 3.78-3.50(m, 4H, N—CH₂ of pyrrolidide), 3.55-3.40 (m, 4H, N—CH₂ of piperazine),2.97 (dd, 1H, J=10.2, 6.6 Hz, H_(5a) of proline), 2.85-2.75 (m, 1H,H_(4b) of proline), 2.69 (dd, 1H, J=10.0, 9.1 Hz, H_(5b) of proline),2.55-2.20 (m, 7H, overlapping N—CH₂ of piperazine, CH₂ of pyrrolidideand H_(3b) of proline), 1.47-1.38 (m, 1H, H_(3a) of proline).

Alternatively, the dihydrochloride salt of the titled compound wasprepared according to the method of Example 1.

Example 114{(2S,4S)-[4-(4-Pyrimidin-2-yl-piperazin-1-yl)-pyrrolidin-2-yl}-(3,3,4,4-tetrafluoro-pyrrolidin-1-yl)-methanonedihydrochloride Step1—tert-Butyl(2S,4S)-4-(4-pyrimidin-2-ylpiperazin-1-yl)-2-[(3,3,4,4-tetrafluoropyrrolidin-1-yl)carbonyl]pyrrolidine-1-carboxylate

DIPEA (261 mL, 1.5 mmol) was added dropwise to a suspension of the titlecompound of Preparation 3 (114 mg, 0.3 mmol), HATU (128 mg, 0.33 mmol),and 3,3,4,4-tetrafluoropyrrolidine hydrochloride (54 mg, 0.3 mmol) in 5mL dichloromethane. After stirring overnight, saturated sodiumbicarbonate solution was added, the mixture was extracted withdichloromethane, the extracts dried over magnesium sulfate, andconcentrated. The residue was purified by chromatography (Biotage® Flash40S, EtOAc) to afford the title compound. MS m/z 503 (MH⁺).

Step 2

An EtOAc/MeOH solution of the product from Step 1 was treated with 4MHCl in dioxane (ca. 5 mL). After 18 hr, the solvent was removed and theresidue was taken up in acetonitrile and concentrated. The solid wastaken up in hexanes, filtered, and dried to afford 50 mg (33%, twosteps) of the title compound. MS m/z 403 (MH⁺).

Using appropriate starting materials, the hydrochloride salts of thecompounds of Examples 115 to 122, disclosed in Table 2, were prepared ina manner analogous to that described in Example 114. TABLE 2 ExampleName MS(M + 1) 115(3-Fluoroazetidin-1-yl)-((2S,4S)-4-(4-(pyrimidin-2-yl)piperazin-1- 335yl)pyrrolidin-2-yl)-methanone 116((3R*,4R*)-3,4-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(pyrimidin-2- 367yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone 117((S)-3-Fluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(pyrimidin-2-yl)piperazin-349 1-yl)pyrrolidin-2-yl)-methanone 118((R)-3-Fluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(pyrimidin-2-yl)piperazin-349 1-yl)pyrrolidin-2-yl)-methanone 119(3,3-Difluoroazetidin-1-yl)((4S)-4-(4-(pyrimidin-2-yl)piperazin-1- 353.3yl)pyrrolidin-2-yl)methanone 120(2S,4S)-4-Fluoro-1-[(2S,4S)-4-(4-pyrimidin-2-yl-piperazin-1-yl)- 374.1pyrrolidine-2-carbonyl]-pyrrolidine-2-carbonitrile 121(S)-4,4-Difluoro-1-[(2S,4S)-4-(2-trifluoromethyl-7,8-dihydro-5H- 431.2pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidine-2-carbonyl]-pyrrolidine-2-carbonitrile 122(2S,4S)-4-Fluoro-1-[(2S,4S)-4-(2-trifluoromethyl-7,8-dihydro-5H- 413.3pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidine-2-carbonyl]-pyrrolidine-2-carbonitrile 123(Azetidin-1-yl)((2S,4S)-4-(4-(pyrimidin-2-yl)piperazin-1-yl)pyrrolidin-317 2-yl)methanone

Example 124((2S,3R,4S)-4-(4-(3-(Trifluoromethyl)pyridin-2-yl)piperazin-1-yl)-3-methylpyrimidin-2-yl)(3,3-difluoropyrrolidin-1-yl)methanonedihydrochloride Step 1

The title compound of Preparation 1 (5.6 g, 20 mmol) was dissolved inbenzene (50 mL) containing 4 Å molecular sieves (7.9 g) and treated withpyrrolidine (2.0 mL, 24 mmol). The solution was filtered andconcentrated to dryness, leaving an orange foam (7.0 g, 100% yield).

Step 2

A solution of the product of Step 1 (7.0 g, 20 mmol) in acetonitrile(100 mL) was added to crushed potassium carbonate (5.2 g, 38 mmol) andtreated with methyl iodide (1.5 mL, 24 mmol). The mixture was heated to90° C. for 16 hers, cooled to RT, and concentrated. The residue wastaken up in chloroform (150 mL) and a mixture of AcOH (5 mL) and water(45 mL) was added. After three hr at RT, the layers were separated, theaqueous layer was extracted with chloroform (3×25 mL), and the combinedorganic phases were washed with saturated sodium bicarbonate (2×25 mL)and brine, and concentrated to a brown oil. The oil was dissolved inether (75 mL), filtered, and concentrated to a pale brown solid (0.97 g,16% yield).

Step 3

To a mixture of the product of Step 2 (74 mg, 0.25 mmol),1-(3-trifluoromethyl)pyridin-2-yl-piperazine (63 mg, 0.28 mol), AcOH (16μL), and sodium acetate (23 mg, 0.28 mmol) in MeOH (1 mL) was addedsodium cyanoborohydride (21 mg, 0.28 mmol). The mixture was stirred atRT for 65 hr and then concentrated. The residue was taken up in EtOAc(20 mL) and the solution was washed with 1 N sodium hydroxide (2×3 mL)and brine (5 mL), dried over magnesium sulfate, and concentrated todryness. The residue was purified by preparative HPLC (Shimadzu,Columbia, Md.; 30×50 cm Waters-Xterra® C18 column—Waters Instrument Co.,Milford, Ma.; 30 mL/min gradient of 15% acetonitrile with 0.1% ammoniumhydroxide over 10 min) to afford a colorless solid (35.7 mg, 26% yield).

Step 4

HCl (4M) in dioxane (0.5 mL) was added to a solution of the product ofStep 3 (35 mg, 0.064 mmol) in acetonitrile (1 mL). After 16 hr, themixture was concentrated to dryness and the residue was triturated withether (2 mL). The title compound was obtained as a solid (32 mg, 96%yield). MS m/z 448.4 (MH⁺).

Using appropriate starting materials, the hydrochloride salts of thecompounds of Examples 125 to 127, disclosed in Table 3 hereinbelow, wereprepared in a manner analogous to that described in Example 124. TABLE 3Example Name MS(M + 1) 125 ((2S,3R,4S)-4-(4-(2-tert-Butyl-5- 544.5(trifluoromethyl)pyrazolo[1,5- a]pyrimidin-7-yl)piperazin-1-yl)-3-methylpyrrolidin-2-yl)(3,3- difluoropyrrolidin-1-yl)methanone 126(3,3-Difluoro-pyrrolidin-1-yl)- 448.4 {(2S,3R,4S)-3-methyl-4-[4-(5-trifluoromethyl-pyridin-2-yl)- piperazin-1-yl]-pyrrolidin-2-yl}-methanone 127 (3,3-Difluoro-pyrrolidin-1-yl)-[(2S,3R,4S)- 381.43-methyl-4-(4-pyrimidin-2- yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone

Example 128(2,4-Difluoro-phenyl)-{4-[(3S,5S)-5-(3,3-difluoro-pyrrolidine-1-carbonyl)-pyrrolidin-3-yl]-piperazin-1-yl}-methanonedihydrochloride Step1—(2S,4S)-4-[4-(2,4-Difluoro-benzoyl)-piperazin-1-yl]-2-(3,3-difluoro-pyrrolidine-1-carbonyl)-pyrrolidine-1-carboxylicacid tert-butyl ester

The title compound of Preparation 7 (97 mg, 0.25 mmol),2,4-difluorobenzoic acid (40 mg, 0.25 mmol and HATU (95 mg, 0.3 mmol)were mixed in anhydrous methylene chloride under nitrogen and cooled to0° C. in and ice bath before addition of DIEA (32 mg, 45 μL, 0.3 mmol).The reaction mixture was allowed to warm to RT and stirred overnight.The reaction was quenched with saturated sodium bicarbonate and theaqueous layer was extracted with methylene chloride. The combinedorganic extracts were washed with brine and dried over magnesiumsulfate. The crude product was purified by flash chromatography usingmethylene chloride-MeOH (95:5) to give the final product as white powder(132 mg, 100%). MS m/z 529.4 (MH⁺). Step 2—An acetonitrile solution ofthe product of Step 1 (120 mg) was treated with 4N HCl in dioxane (1mL). The reaction was stirred at RT overnight and evaporated. Theresidue was dissolved in water, filtered, and lyophilized overnight toafford the title product as white powder (110 mg, 96%). MS m/z 429.2(MH⁺).

Using appropriate starting materials, the hydrochloride salts of thecompounds of Examples 129 to 133, disclosed in Table 4, were prepared ina manner analogous to that described in Example 128. TABLE 4 ExampleName MS(M + 1) 129(3,3-Difluoro-pyrrolidin-1-yl)-{(2S,4S)-4-[4-(toluene-4-sulfonyl)- 443.2piperazin-1-yl]-pyrrolidin-2-yl}-methanone 130(3-Amino-pyrazin-2-yl)-{4-[(3S,5S)-5-(3,3-difluoro-pyrrolidine-1- 410.2carbonyl)-pyrrolidin-3-yl]-piperazin-1-yl}-methanone 131{4-[(3S,5S)-5-(3,3-Difluoro-pyrrolidine-1-carbonyl)-pyrrolidin-3-yl]-444.3 piperazin-1-yl}-quinolin-4-yl-methanone 1324-[(3S,5S)-5-(3,3-Difluoro-pyrrolidine-1-carbonyl)-pyrrolidin-3-yl]-360.2 piperazine-1-carboxylic acid-ethylamide 1334-[(3S,5S)-5-(3,3-Difluoro-pyrrolidine-1-carbonyl)-pyrrolidin-3-yl]-426.2 piperazine-1-carboxylic acid-(4-fluoro-phenyl)-amide

BIOLOGICAL METHODOLOGIES

The utility of the compounds of formula (I), the prodrugs andstereoisomers thereof, and the pharmaceutically acceptable salts of thecompounds, prodrugs, and stereoisomers, in the treatment or preventionof the conditions enumerated hereinabove in mammals may be demonstratedin conventional assays known to one of ordinary skill in the relevantart, including the in vivo and in vitro assays described below. Suchassays also provide a means by which the activities of the compounds offormula (I), the prodrugs, and stereoisomers thereof, and thepharmaceutically acceptable salts of the compounds, prodrugs, andstereoisomers, may be compared with the activities of other compounds.

In Vitro Assay for DPP-IV Inhibition

DPP-IV inhibition may be demonstrated in vitro by the following assay,which is adapted from methods of Scharpe, et al., A. Clin. Chem., 2299(1988) and Lodja, Z. Czechoslovak Medicine, 181 (1988). 150 μL of anenzyme-substrate solution is pipetted into microtiter wells of apolystyrene 96-well plate, and maintained at 4° C. The enzyme-substratesolution comprises 50 μM Gly-Pro-4-methoxy-β-naphthylamide hydrochloridein 50 mM Tris assay buffer pH 7.3 containing 0.1M sodium chloride, 0.1%(v/v) Triton and 50 μU/mL DPP-IV (MP Biomedicals, Livermore, Calif.;DPP-IV 5 mU/mL stock). 5 μL per well of the compound of formula (I) isadded, bringing the final concentrations of the formula (I) compound tobetween 3 μM and 10 nM per well.

Controls. Enzyme is omitted from four (4) wells, as a reagent blank. 5μL of 3 mM Diprotin A (Bachem Bioscience, Inc.; King of Prussia, Pa.) isadded to four wells as a positive quality control, providing a finalDiprotin A concentration of 100 μM. To measure total enzyme activity(i.e., a negative control), without the influence of any compounds offormula (I), 5 μL of distilled water is added to four wells.

The entire assay is incubated overnight (between 14 and 18 hours) at 37°C. The reaction is quenched by adding 10 μL of Fast Blue B solution (0.5mg/mL Fast Blue B in a buffer comprising 0.1M sodium acetate pH 4.2 and10% (v/v) Triton X-100 to each well, followed by shaking forapproximately 5 min at room temperature. The plates may be analyzed on aSpectramax spectrophotometer (Molecular Devices; Sunnyvale, Calif.), orequivalent equipment, (absorption maximum at 525 nm). IC₅₀ data forcompounds may be obtained by measuring the activity of DPP-IV over arange of compound concentrations from 10 nM to 3 μM.

In Vivo Assay for Glucose Lowering

The glucose lowering effects of DPP-IV inhibitors, including thecompounds of formula (I), may be exemplified in 4-6 week old KK/H1J mice(Jackson Labs; Bar Harbor, Me.) in the context of an oral glucosetolerance test.

Oral glucose tolerance tests (OGTT) have been in use in humans since, atleast, the 1930s, as described by Pincus, et al., Am. J. Med. Sci., 782(1934), and are routinely used in the diagnosis of human diabetes,though not to evaluate the efficacy of therapeutic agents in patients.

KK mice have been used to evaluate (i) glitazones (Fujita et al.Diabetes, 804 (1983); Fujiwara, et al., Diabetes, 1549 (1988); andIzumi, et al., Biopharm Drug. Dispos., 247 (1997)); (ii) metformin(Reddi, et al., Diabet. Metabol., 44 (1993)); (iii) glucosidaseinhibitors (Hamada, et al., Jap. Pharmacol. Ther., 17 (1988) and Matsuo,et al., Am. J. Clin. Nutr., 314S (1992)), and (iv) extra-pancreaticeffects of sulfonylureas (Kameda, et al., Arzneim. Forsch./Drug Res.,39044 (1982) and Muller et al., Horm. Metabl. Res., 469 (1990)).

KK mice are derived from an inbred line first established and describedby Kondo, et al., Bull. Exp. Anim., 107 (1957). These mice spontaneouslydevelop a hereditary form of polygenic diabetes that progresses to causerenal, retinal, and neurological complications analogous to those seenin human diabetic subjects, however, they do not require insulin orother medication for survival.

Another aspect of the invention is directed to the use of KK mice toevaluate the effects of insulin secretagogue agents in the context of anoral glucose tolerance test. The mice are fasted overnight (about 14 toabout 18 hr), but allowed free access to water. After fasting, (time“t=0), 25 μL of blood is drawn from the retro-orbital sinus and added to0.025% heparinized saline (100 μL) on ice. The mice (10 per group) arethen orally dosed with a solution of a compound of formula (I) in 0.5%methylcellulose (0.2 mL/mouse). Two controls groups receive only 0.5%methylcellulose. At t=15 min, the mice are bled, as described above, andthen dosed with 1 mg/kg glucose in distilled water (0.2 mL/mouse). Thefirst control group is dosed with glucose. The second control group isdosed with water. At t=45 min, the mice are again bled, as describedabove. The blood samples are centrifuged, the plasma collected andanalyzed for glucose content on a Roche-Hitachi 912 glucose analyzer(Roche Diagnostics Corp.; Indianapolis, Ind.). The data may be expressedas percent (%) inhibition of glucose excursion relative to the twocontrol groups (i.e., the glucose level in the animals receiving glucosebut no test compound representing 0% inhibition and the glucoseconcentration in the animals receiving only water representing 100%inhibition).

The compounds of formula (I) generally exhibit inhibitory activity,expressed as IC₅₀'s, against DPP-IV that are <1,000 nM. Generallypreferred compounds have IC₅₀'s<100 nM. For example,((2S,4S)-4-(4-(3-cyanopyrazin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)-((3R*,4S)-3,4-difluoropyrrolidin-1-yl)-methanonedihydrochloride has an IC₅₀ of 3.5 nM.

Comparative Rat Pharmacokinetics Experiments

Rat Pharmacokinetics experiments were performed to demonstrate theimprovement in plasma concentrations maintained over time for a compoundof the present invention as compared to a structurally similar prior artcompound generically disclosed in International Application WO 02/14271.Specifically, plasma concentrations over time were measured for ratsadministered (a) the dihydrochloride salt of(3,3-difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(pyrimidin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone(hereinafter “CPD 113”), which was prepared as described in Example 113,and (b) the comparative dihydrochloride salt of((2S,4S)-4-(4-(pyrimidin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)(pyrrolidin-1-yl)methanone(hereinafter “comparator”), which may be prepared according to themethod of Example 1 or as generally described in WO 02/14271.

In this experiment, male Sprague-Dawley rats (200-250 grams) implantedwith jugular vein cannulas (JVC) were obtained from Charles RiverLaboratories. Each compound was administered to two rats or by oralgavage. The oral dose was administered as a solution in 0.5%methycellulose with a dose volume of 10 mL/kg. The amount of eachcompound administered was 5 mg/kg body weight. Blood samples (0.25 mL)were collected at multiple time points from 0-24 hours and placed intotubes containing lithium heparin (Becton Dickinson, Microtainer®). Theblood samples were then centrifuged at 12000 rpm for 10 minutes). Plasmaaliquots were taken for determination of compound plasma concentrations(pharmacokinetic analysis). The plasma samples were frozen at −70° C.until analysis.

The rat plasma samples were analyzed for compound concentrations byLC/MS/MS (Applied Biosystems API 40000 mass spectrometer). In brief,compound standard curves were prepared in control rat plasma with adynamic range of 1.0-2000 ng/mL. Aliquots (0.02 mL) of both standardsand samples were placed into Marsh® tubes in a 96-well block. Proteinswere precipitated by addition of 0.1 mL acetonitrile containing 0.1μg/mL of internal standard. The 96-well blocks were vortexed and thencentrifuged at 3000 rpm for 5 minutes. The resulting supernatant wasremoved and placed into a new 96-well block and taken to dryness at 50°C. under a nitrogen stream. Residues were reconstituted in mobile phase(60% 5 mM ammonium acetate and 40% acetonitrile). Aliquots (0.01 mL)were then injected onto the LC/MS/MS for analysis.

The average plasma concentrations, measured are provided in thefollowing table. CPD 113 Comparator Mean Mean Plasma Plasma Compound/Level Std Level Std Time (hr) ng/ml dev ng/ml dev 0.25 1406.0 338.0 44671.1 0.5 1322.5 359.9 425 108 0.75 979.2 137.0 319 59.8 1 768.2 314.0283 13.3 2 289.2 71.8 128 40.4 4 97.8 69.2 27.3 11.2 6 49.3 19.1 12.71.2 8 32.8 25.5 6.16 2.62

As shown by their despective plasma concentrations, CPD 113 achieved andmaintained significantly higher plasma concentrations than did thecomparator compound.

1. A compound of formula (I)

or a pharmaceutically acceptable salt of said compound, or a solvate ofsaid compound or salt, wherein: R¹ is —(C₁-C₆)alkyl, —(C₁-C₆)alkoxy,—(C₁-C₆)arylalkyl, —NR^(a)R^(b), hydroxy, cyano, aryl, or heteroaryl,wherein said —(C₁-C₆)alkyl, said aryl, or said heteroaryl is optionallysubstituted independently with one to three —COOH, —C(O)(C₁-C₆)alkoxy,—C(O)(C₁-C₆)alkyl, —C(O)NR^(a)R^(b), cyano, halogen, nitro,trifluoromethyl, —(C₁-C₆)alkyl, —(C₁-C₆)alkoxy, —(C₃-C₆)cycloalkyl, orphenyl, wherein: R^(a) and R^(b) are, independently, hydrogen,—(C₁-C₆)alkyl, aryl, or heteroaryl, or R^(a) and R^(b), taken togetherwith the nitrogen atom to which they are attached, form a four- tosix-membered heterocyclic ring, wherein said ring optionallyincorporates an additional one or two nitrogen, oxygen, or sulfur ringheteroatoms; R² and R³ are, independently, hydrogen, halogen,—(C₁-C₆)alkyl, or —(C₃-C₈)cycloalkyl; Q is a covalent bond, —C(O)—, or—SO₂—; HET is a heterocycloalkyl ring moiety, optionally substitutedwith: (A) one to four —(C₁-C₆)alkyl, optionally substituted with one tosix halogen atoms, —(C₁-C₆)alkoxy, cyano, halogen, hydroxy, or—NR^(a)R^(b), or (B) —(C₁-C₆)arylalkyl, optionally substituted with oneto six halogen atoms, —(C₁-C₆)alkoxy, cyano, halogen, hydroxy, or—NR^(a)R^(b); n is 0 or 1; when n is 0, X is —CH₂—, and Y is —CH₂—,—CHF—, or —CF₂—; or when n is 1, X is —CH₂—, —CHF—, or —CF₂ 13 ; and Yis —CH₂—, —CHF—, or —CF₂—, provided that X and Y are not both —CH₂—; andZ is hydrogen or cyano.
 2. The compound of claim 1, wherein: R¹ is arylor heteroaryl, optionally substituted independently with one to threecyano, halogen, nitro, trifluoromethyl, —(C₁-C₆)alkyl, —(C₁-C₆)alkoxy,—(C₃-C₆)cycloalkyl, or phenyl; R² is —H or —(C₁-C₆)alkyl; R³ is —H or—(C₁-C₆)alkyl; and HET is azetidinyl, piperazinyl, piperidinyl,pyrrolidinyl, 5,6-dihydro-8H-imidazo[1,2-a]pyrazin-7-yl,5,6-dihydro-8H-[1,2,4]triazolo[4,3-a]pyrazin-7-yl, or7,8-dihydro-5H-pyrido[4,3-a]pyrimidin-6-yl.
 3. The compound of claim 1,wherein: R¹ is benzoisothiazolyl, benzisoxazolyl, isothiazolyl,isoxazolyl, oxazolopyridyl, pyrazinyl, pyridinyl, pyrimidinyl,quinolinyl, quinoxalinyl, thiadiazolyl, triazinyl, or1,1-dioxo-1H-1,2-benzoisothiazolyl; R² and R³ are —H; Q is a covalentbond; and HET is piperazinyl.
 4. The compound of claim 3 wherein R¹ ispyridinyl or pyrimidinyl.
 5. The compound of claim 4 wherein n is 1, Xis —CF₂— and Y is —CH₂—.
 6. The compound of claim 1 selected from thegroup consisting of((2S,4S)-4-(3-(trifluoromethyl)-5,6-dihydro-[1,2,4]triazolo[4,3-a]pyrazin-7(8H)-yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone,(3,3-difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(oxazolo[5,4-b]pyridin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone,(3,3-difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(4-methylpyrimidin-2-yl)piperazin-1-yl)pyrrolidin-2-yl)-methanone,((2S,4S)-4-(2-(trifluoromethy)-7,8-dihydropyrido[4,3-d]pyrimidin-6(5H)-yl)pyrrolidin-2-yl)-(3,3-difluoropyrrolidin-1-yl)-methanone,((S)-3-fluoro-pyrrolidin-1-yl)-{(2S,4S)-4-[4-(3-trifluoromethyl-pyridin-2-yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone,((S)-3-fluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-trifluoromethyl-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone,(3,3-difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[4,5-c]pyridin-2-yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone,[(2S,4S)-4-(2-cyclopropyl-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-(3-fluoro-azetidin-1-yl)-methanone,(3,3-difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-ethoxy-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone,2-{4-[(3S,5S)-5-(3-fluoro-azetidine-1-carbonyl)-pyrrolidin-3-yl]-piperazin-1-yl}-nicotinonitrile,((S)-3-fluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-b]pyridin-2-yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone,(3-fluoro-azetidin-1-yl)-[(2S,4S)-4-(2-trifluoromethyl-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone,2-{4-[(3S,5S)-5-((S)-3-fluoro-pyrrolidine-1-carbonyl)-pyrrolidin-3-yl]-piperazin-1-yl}-nicotinonitrile,(3-fluoro-azetidin-1-yl)-{(2S,4S)-4-[4-(2-trifluoromethyl-quinolin-4-yl)-piperazin-1-yl]-pyrrolidin-2-yl}-methanone,((3R*,4S*)-3,4-difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(2-trifluoromethyl-7,8-dihydro-5H-pyrido[4,3-d]pyrimidin-6-yl)-pyrrolidin-2-yl]-methanone,and((3R*,4S*)-3,4-difluoro-pyrrolidin-1-yl)-[(2S,4S)-4-(4-oxazolo[5,4-b]pyridin-2yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone;or a pharmaceutically acceptable salt of said compound.
 7. A compound ofclaim 1, 4 or 5, or a pharmaceutically acceptable salt of said compound,or a solvate of said compound or salt, for use in therapy.
 8. Apharmaceutical composition comprising: (a) a compound of claim 1, 4 or5, or a pharmaceutically acceptable salt of said compound, or a solvateof said compound or salt; and (b) a pharmaceutically acceptable carrier,vehicle, diluent or excipient.
 9. A method of inhibiting dipeptidylpeptidase-IV in a mammal comprising administering to said mammal in needof such treatment a therapeutically effective amount of a compound ofclaim 1, 4 or 5, or a pharmaceutically acceptable salt of said compound,or a solvate of said compound or salt.
 10. A method of treating Type 2diabetes in a mammal in need of such treatment a therapeuticallyeffective amount of a compound of claim 1, 4 or 5, or a pharmaceuticallyacceptable salt of said compound, or a solvate of said compound or salt.11. A method of treating Type 1 diabetes in a mammal in need of suchtreatment a therapeutically effective amount of a compound of claim 1, 4or 5, or a pharmaceutically acceptable salt of said compound, or asolvate of said compound or salt.
 12. A method of treating hyperglycemiain a mammal in need of such treatment a therapeutically effective amountof a compound of claim 1, 4 or 5, or a pharmaceutically acceptable saltof said compound, or a solvate of said compound or salt.
 13. A method oftreating metabolic syndrome in a mammal in need of such treatment atherapeutically effective amount of a compound of claim 1, 4 or 5, or apharmaceutically acceptable salt of said compound, or a solvate of saidcompound or salt.
 14. A method of treating impaired glucose tolerance ina mammal in need of such treatment a therapeutically effective amount ofa compound of claim 1, 4 or 5, or a pharmaceutically acceptable salt ofsaid compound, or a solvate of said compound or salt.
 15. A method oftreating diabetic neuropathy in a mammal in need of such treatment atherapeutically effective amount of a compound of claim 1, 4 or 5, or apharmaceutically acceptable salt of said compound, or a solvate of saidcompound or salt.
 16. A method of treating diabetic nephropathy in amammal in need of such treatment a therapeutically effective amount of acompound of claim 1, 4 or 5, or a pharmaceutically acceptable salt ofsaid compound, or a solvate of said compound or salt.
 17. A method oftreating diabetic retinopathy in a mammal in need of such treatment atherapeutically effective amount of a compound of claim 1, 4 or 5, or apharmaceutically acceptable salt of said compound, or a solvate of saidcompound or salt.
 18. A method of treating diabetic cardiomyopathy in amammal in need of such treatment a therapeutically effective amount of acompound of claim 1, 4 or 5, or a pharmaceutically acceptable salt ofsaid compound, or a solvate of said compound or salt.
 19. The method ofclaim 10 wherein said mammal is a human.